Team:UT-Tokyo/LabNote
From 2011.igem.org
(Difference between revisions)
(46 intermediate revisions not shown) | |||
Line 2: | Line 2: | ||
=Parts List= | =Parts List= | ||
+ | |||
+ | Each part is represented by s number (e.g. #20 for lac promotor) in this notebook. | ||
+ | By pointing the mouse on the number within notebook entires, you can find out the details of the part. | ||
+ | |||
+ | |||
{| id="partslist" | {| id="partslist" | ||
- | |+ Table 1. List of iGEM parts | + | |+ Table 1. List of iGEM parts used in our project |
|- | |- | ||
- | ! Number !! | + | ! Number !! Reg.ID !! Content !! Plasmid !! Length (bp) |
|- | |- | ||
- | | #1 || J23119 || c.Promoter (Strong) || pSB1A2 || class="number" | 35 | + | | #1 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23119 J23119] || c.Promoter (Strong) || pSB1A2 || class="number" | 35 |
|- | |- | ||
- | | #2 || J23118 || c.Promoter (Medium) || BBa_J61002 || class="number" |35 | + | | #2 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23118 J23118] || c.Promoter (Medium) || [http://partsregistry.org/wiki/index.php?title=Part:BBa_J61002 BBa_J61002] || class="number" |35 |
|- | |- | ||
- | | #3 || B0032 || RBS || pSB1A2 || class="number" |13 | + | | #3 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_B0032 B0032] || RBS || pSB1A2 || class="number" |13 |
|- | |- | ||
- | | #5 || B0014 || d.Ter || pSB1AK3 || class="number" |95 | + | | #5 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_B0014 B0014] || d.Ter || pSB1AK3 || class="number" |95 |
|- | |- | ||
- | | #9 || E0240 || RBS-GFP-d.Ter || pSB1A2 || class="number" |876 | + | | #9 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_E0240 E0240] || RBS-GFP-d.Ter || pSB1A2 || class="number" |876 |
|- | |- | ||
- | | #10 || E0040 || GFP || pSB1A2 || class="number" |681 | + | | #10 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_E0040 E0040] || GFP || pSB1A2 || class="number" |681 |
|- | |- | ||
- | | #11 || E1010 || RFP || pSB2K3 || class="number" |723 | + | | #11 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_E1010 E1010] || RFP || pSB2K3 || class="number" |723 |
|- | |- | ||
- | | #14 || I712019 || fLuc || pSB1AK8 || class="number" |1653 | + | | #14 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_I712019 I712019] || fLuc || pSB1AK8 || class="number" |1653 |
|- | |- | ||
- | | #17 || J52008 || rLuc || pSB1AK3 || class="number" |936 | + | | #17 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_J52008 J52008] || rLuc || pSB1AK3 || class="number" |936 |
|- | |- | ||
- | | #20 || R0011 || lacP || pSB1A2 || class="number" |55 | + | | #20 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0011 R0011] || lacP || pSB1A2 || class="number" |55 |
|- | |- | ||
- | | #21 || C0012 || LacI || pSB1A2 || class="number" |1128 | + | | #21 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0012 C0012] || LacI || pSB1A2 || class="number" |1128 |
|- | |- | ||
- | | #22 || I712074 || pT7 || pSB1AK8 || class="number" |46 | + | | #22 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_I712074 I712074] || pT7 || pSB1AK8 || class="number" |46 |
|- | |- | ||
- | | #23 || K145001 || T7 RNA Pol. || pSB1A2 || class="number" |2655 | + | | #23 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_K145001 K145001] || T7 RNA Pol. || pSB1A2 || class="number" |2655 |
|- | |- | ||
- | | #24 || J22106 || recAp || pSB1A2 || class="number" |192 | + | | #24 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_J22106 J22106] || recAp || pSB1A2 || class="number" |192 |
|- | |- | ||
- | | #27 || C0083 || AspA || pSB2K3 || class="number" |1518 | + | | #27 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0083 C0083] || AspA || pSB2K3 || class="number" |1518 |
|- | |- | ||
- | | #28 || K112808 || T4 phage lysis device (no promoter) || pSB1A2 || class="number" | 1785 | + | | #28 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_K112808 K112808] || T4 phage lysis device (no promoter) || pSB1A2 || class="number" | 1785 |
|- | |- | ||
| #29 || - || CheZ || pSB1AK3 || class="number" | 728 | | #29 || - || CheZ || pSB1AK3 || class="number" | 728 | ||
|- | |- | ||
- | | #30 || K117000 || Lysis gene || pSB1A2 || class="number" | 144 | + | | #30 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_K117000 K117000] || Lysis gene || pSB1A2 || class="number" | 144 |
|- | |- | ||
| #31 || - || LexA || pSB1AK3 || class="number" | 750 | | #31 || - || LexA || pSB1AK3 || class="number" | 750 | ||
Line 51: | Line 56: | ||
| #35 || - || recNp || || | | #35 || - || recNp || || | ||
|- | |- | ||
- | | #36 || R0051 || cI-repressed promoter || pSB1A2 || class="number" | 49 | + | | #36 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0051 R0051] || cI-repressed promoter || pSB1A2 || class="number" | 49 |
|- | |- | ||
- | | #37 || C0051 || cI repressor (LVA tagged) || pSB1A2 || \ | + | | #37 || [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0051 C0051] || cI repressor (LVA tagged) || pSB1A2 || \ |
|- | |- | ||
| #38 || - || RecA || || | | #38 || - || RecA || || | ||
Line 149: | Line 154: | ||
--> | --> | ||
- | =Lab | + | =Lab Notebook= |
- | + | ||
- | + | ||
<html> | <html> | ||
+ | |||
+ | <ul id="months"> | ||
+ | <li>May</li> | ||
+ | <li>June</li> | ||
+ | <li>July</li> | ||
+ | <li>August</li> | ||
+ | <li>September</li> | ||
+ | <li>October</li> | ||
+ | </ul><br /> | ||
+ | |||
+ | <div id="calendar"> | ||
+ | <p>Please enable Javascript to view this calendar.</p> | ||
+ | </div> | ||
+ | |||
<div id="labnote"> | <div id="labnote"> | ||
</html> | </html> | ||
- | =='11/9/12 (Mon)== | + | <html><div class="labMonth" id="labMonth_5"></html> |
+ | ==May== | ||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/05/18 (Wed)=== | ||
+ | *Making LB medium, 50×TAE, Tris-HCl (pH8.0) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/05/24 (Tue)=== | ||
+ | *Making SOB medium, 0.5M EDTA (pH8.0) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/05/25(Wed)=== | ||
+ | *Making TB(pH=6.7), LB plate | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/05/26(Thu)=== | ||
+ | *Making Mgaq(for SOB medium), Competent cell | ||
+ | *TB filtration | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/05/31(Tue)=== | ||
+ | *iGEM parts resuspension + frozen stock (at -20℃) | ||
+ | *Transformation | ||
+ | *Overnight culture on LB plate (with 100ug/ml ampicilline) | ||
+ | <html></div></html> | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labMonth" id="labMonth_6"></html> | ||
+ | ==June== | ||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/01(Wed)=== | ||
+ | *Picking up colony and transfer to LB medium | ||
+ | *Making LB medium | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/02(Thu)=== | ||
+ | *Miniprep | ||
+ | *Making Glycerol(50%)(for cryopreservation) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/07(Tue)=== | ||
+ | *Nanodrop | ||
+ | *Transformation | ||
+ | *Culture from frozen stock | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/08(Wed)=== | ||
+ | *Picking up colony | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/09(Thu)=== | ||
+ | *Miniprep | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/13(Mon)=== | ||
+ | *Planting Negative control | ||
+ | *Making frozen stock | ||
+ | *Transformation(BBa_E0240, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, B0015, <nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">10</span></html>, ,J31000, ,J44000) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/14(Tue)=== | ||
+ | *Picking up colony | ||
+ | *Making Mg reagent | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/15(Wed)=== | ||
+ | *Miniprep | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/17(Fri)=== | ||
+ | *Picking up colony(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, B0015, B0040,<nowiki>#</nowiki><html><span class="part_num">9</span></html>, J31000, J44000) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Dissolution(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, K16500, <nowiki>#</nowiki><html><span class="part_num">24</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/21(Tue)=== | ||
+ | *Making frozen stock(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, B0015, B0040, <nowiki>#</nowiki><html><span class="part_num">9</span></html>, J31000, J44000) | ||
+ | *Passafe culture(<nowiki>#</nowiki><html><span class="part_num">9</span></html>, J31000) | ||
+ | *Nanodrop again | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/22(Wed)=== | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">9</span></html>, J31000) | ||
+ | *Making competent cell | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/24(Fri)=== | ||
+ | *Digest (product of Miniprep 06/09 EScut) | ||
+ | *Making agarose gel | ||
+ | *Electrophoresis | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/28(Tue)=== | ||
+ | *Digest(product of Miniprep 06/09 EPcut) | ||
+ | *Electrophoresis | ||
+ | *Defrost and transformation(BBa_E0030, E0020, <nowiki>#</nowiki><html><span class="part_num">14</span></html>, I712052,<nowiki>#</nowiki><html><span class="part_num">17</span></html>) | ||
+ | *Making 1×TAE, agarose gel, LB medium, LB plate(amp), | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/06/29(Wed)=== | ||
+ | *Picking up colony | ||
+ | *Making LB broth | ||
+ | <html></div></html> | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labMonth" id="labMonth_7"></html> | ||
+ | ==July== | ||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/01(Fri)=== | ||
+ | *Digest(EPcut) | ||
+ | *Miniprep(BBa_E0030, E0020, <nowiki>#</nowiki><html><span class="part_num">14</span></html>, I712052, <nowiki>#</nowiki><html><span class="part_num">17</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/05(Tue)=== | ||
+ | *Electrophoresis(product of digest 07/01) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Making antibiotic stock(1000×)(Km,Cm) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/06(Wed)=== | ||
+ | *Gel extraction (pre)(BBa_I712052) | ||
+ | *Picking up colony(Ba_J23119, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span></html>, <nowiki>#</nowiki><html><span class="part_num">10</span></html>) | ||
+ | *Making agarose gel | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/07(Thu)=== | ||
+ | *Digest(BBa_E0240, <nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Defrost(BBa_E1010, K325101, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">21</span></html>, <nowiki>#</nowiki><html><span class="part_num">22</span></html>, <nowiki>#</nowiki><html><span class="part_num">23</span></html>) | ||
+ | *Transformation(BBa_K145001, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">21</span></html>, <nowiki>#</nowiki><html><span class="part_num">22</span></html>) | ||
+ | *Making LB plate(Cm,Km) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/08(Fri)=== | ||
+ | *Miniprep(BBa_J23119, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">10</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/11(Mon)=== | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">9</span></html> <nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span></html>) | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">21</span></html>, K325101, <nowiki>#</nowiki><html><span class="part_num">22</span></html>, <nowiki>#</nowiki><html><span class="part_num">23</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/12(Tue)=== | ||
+ | *Picking up colony | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/13(Wed)=== | ||
+ | *Frozen stock(<nowiki>#</nowiki><html><span class="part_num">1</span></html>(18A), <nowiki>#</nowiki><html><span class="part_num">20</span></html>(6G), <nowiki>#</nowiki><html><span class="part_num">21</span></html>(2O), <nowiki>#</nowiki><html><span class="part_num">14</span></html>(6N), <nowiki>#</nowiki><html><span class="part_num">23</span></html>(2F)) | ||
+ | *Picking up colony(<nowiki>#</nowiki><html><span class="part_num">11</span></html>) | ||
+ | *Gel extraction(B<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Digest (<nowiki>#</nowiki><html><span class="part_num">2</span></html> SPcut) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/14(Thu)=== | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">21</span></html>, <nowiki>#</nowiki><html><span class="part_num">22</span></html>, <nowiki>#</nowiki><html><span class="part_num">23</span></html>) | ||
+ | *Electrophoresis(<nowiki>#</nowiki><html><span class="part_num">2</span></html>) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Passafe(<nowiki>#</nowiki><html><span class="part_num">11</span></html>) | ||
+ | *Making LB+amp | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/15(Fri)=== | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Dispensing Ligation Buffer | ||
+ | *Frozen stock(<nowiki>#</nowiki><html><span class="part_num">11</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/19(Tue)=== | ||
+ | *Digest | ||
+ | *Gel extraction | ||
+ | *Making competent cell | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/20(Wed)=== | ||
+ | *Picking up colony | ||
+ | *Making Master plate | ||
+ | *Testing product of Miniprep | ||
+ | *Transformation (<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/21(Thu)=== | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>) | ||
+ | *Testing product | ||
+ | *Digest | ||
+ | **SPcut : <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span></html> | ||
+ | **XPcut : <nowiki>#</nowiki><html><span class="part_num">10</span></html>, <nowiki>#</nowiki><html><span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | *Making TB | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/22(Fri)=== | ||
+ | *Frozen stock(<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Electrophoresis(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">10</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">11</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">17</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *Digest | ||
+ | **SPcut : <nowiki>#</nowiki><html><span class="part_num">20</span></html> | ||
+ | **XPcut : <nowiki>#</nowiki><html><span class="part_num">10</span></html>, <nowiki>#</nowiki><html><span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>, product of Miniprep 07/20) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/25(Mon)=== | ||
+ | *Colony check(<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>) | ||
+ | *Gel Extraction(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">10</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">11</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">17</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *Ligation and Transformation(<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Defrost Primer(200×, 10×) | ||
+ | *Dispensing PCR Mix | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/26(Tue)=== | ||
+ | *Picking up colony and making master plate(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html> XPcut,Ecut, <nowiki>#</nowiki><html><span class="part_num">22</span></html> SPcut, <nowiki>#</nowiki><html><span class="part_num">23</span></html> XPcut) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(Re), <nowiki>#</nowiki><html><span class="part_num">3</span></html>(Negative control)) | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">15</span></html>, <nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">30</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>(Re), <nowiki>#</nowiki><html><span class="part_num">3</span></html>(Negative control)) | ||
+ | *Making reagent for TB(KOH, CaCl2, KCl, MnCl2) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/27(Wed)=== | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">10</span></html>, <nowiki>#</nowiki><html><span class="part_num">22</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Electrophoresis | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/28(Thu)=== | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">22</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">23</span></html>_XP) | ||
+ | *Digest | ||
+ | **SPcut : <nowiki>#</nowiki><html><span class="part_num">1</span></html>,<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span></html> | ||
+ | **EScut : <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span></html> | ||
+ | **XPcut : <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | *Making TB bugger, LB plate, 0.3% LB plate | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/07/29(Fri)=== | ||
+ | *Cloning(lexA, cheZ) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">1</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">2</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">3</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">24</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP, lexA, cheZ PCR product) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">30</span></html>) | ||
+ | *Digest(lexA, cheZ PCR product XP cut) | ||
+ | <html></div></html> | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labMonth" id="labMonth_8"></html> | ||
+ | ==August== | ||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/01(Mon)=== | ||
+ | *Nanodrop(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(4)M<nowiki>#</nowiki><html><span class="part_num">30</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">3</span></html> SPcut, <nowiki>#</nowiki><html><span class="part_num">5</span></html> EXcut, <nowiki>#</nowiki><html><span class="part_num">9</span></html> XPcut, <nowiki>#</nowiki><html><span class="part_num">30</span></html> XP cut) | ||
+ | *Cloning(lexA, cheZ) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">30</span></html>, pSB1A2) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">23</span></html>, <nowiki>#</nowiki><html><span class="part_num">22</span>-</html><nowiki>#</nowiki><html><span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">30</span></html>) | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">23</span></html>, <nowiki>#</nowiki><html><span class="part_num">22</span>-</html><nowiki>#</nowiki><html><span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">30</span></html>, <nowiki>#</nowiki><html><span class="part_num">27</span></html>(IPTG), <nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/02(Tue)=== | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">23</span></html>, <nowiki>#</nowiki><html><span class="part_num">22</span>-</html><nowiki>#</nowiki><html><span class="part_num">9</span></html>) | ||
+ | *Cloning(PCR)(<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | *Gel extraction | ||
+ | *PCR_2nd(lexA, cheZ, <nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | *Digest (<nowiki>#</nowiki><html><span class="part_num">30</span></html> again) | ||
+ | *Making competent cell(cheZ-, Wild Type) | ||
+ | *Miniprep | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/04(Thu)=== | ||
+ | *Electrophoresis(product of ligation <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1,2), <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">22</span>-<span class="part_num">9</span></html>(1,2,3)) | ||
+ | *Frozen stock(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1,2), <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">22</span>-<span class="part_num">9</span></html>(1,2)) | ||
+ | *Making Gel | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">23</span></html> XPcut) | ||
+ | *IPTG induct(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">9</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/05(Fri)=== | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(2) XPcut, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1) XPcut) | ||
+ | *Cloning(PCR)(<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>, lexA, cheZ) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/08(Mon)=== | ||
+ | *Defrost Frozen stock, Miniprep, Nanodrop and Electrophoresis(<nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">23</span></html>, <nowiki>#</nowiki><html><span class="part_num">30</span></html>) | ||
+ | *Making Gel | ||
+ | *Digest | ||
+ | **XPcut : <nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>, cheZ, lexA, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">23</span></html>,<nowiki>#</nowiki><html><span class="part_num">30</span></html> | ||
+ | **EScut : <nowiki>#</nowiki><html><span class="part_num">6</span></html> | ||
+ | **EXcut : <nowiki>#</nowiki><html><span class="part_num">7</span></html> | ||
+ | **SPcut : <nowiki>#</nowiki><html><span class="part_num">7</span></html>,<nowiki>#</nowiki><html><span class="part_num">20</span></html> | ||
+ | *Defrost Frozen stock(<nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1)) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/09(Tue)=== | ||
+ | *Miniprep, Electrophoresis and Digest(<nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1)) | ||
+ | *Gel extraction and Nanodrop(<nowiki>#</nowiki><html><span class="part_num">6</span></html>, <nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">8</span></html>, cheZ, lexA③, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">23</span></html>, <nowiki>#</nowiki><html><span class="part_num">7</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">7</span></html>_SP) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">27</span>-</html>psB1A2, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span>-</html>psB1A2, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">29</span>-</html>psB1A2, <nowiki>#</nowiki><html><span class="part_num">31</span>-</html>psB1A2, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">23</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">30</span></html>) | ||
+ | *Making LB plate | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/10(Wed)=== | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">27</span>-</html>pSB1A2, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span>-</html>psB1A2, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">29</span>-</html>psB1A2, <nowiki>#</nowiki><html><span class="part_num">31</span>-</html>pSB1A2, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">23</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(1), <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">30</span></html>)) | ||
+ | *Cloning(lexA, cheZ) | ||
+ | *Digest | ||
+ | **EScut : <nowiki>#</nowiki><html><span class="part_num">6</span></html> | ||
+ | **EXcut : <nowiki>#</nowiki><html><span class="part_num">7</span></html> | ||
+ | **SPcut : <nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">7</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html> | ||
+ | **XPcut : <nowiki>#</nowiki><html><span class="part_num">23</span></html> | ||
+ | *Defrost Primer | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/11(Thu)=== | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">30</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(insert,vector), <nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, cheZ, lexA) | ||
+ | *Cloning(sulAp, uvrAp) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">31</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/12(Fri)=== | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">6</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">23</span></html>) | ||
+ | *Cloning(nested-PCR)(uvrAp, sulAp) | ||
+ | *Check Electrophoresis | ||
+ | **8/11 PCR product:<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html> | ||
+ | **Miniprep product:<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">5</span></html> | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">5</span></html> EXcut, <nowiki>#</nowiki><html><span class="part_num">14</span></html> EScut) | ||
+ | *Gel extraction | ||
+ | **8/11 digest product : <nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>, cheZ, lexA | ||
+ | **8/11 PCR product : <nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html> | ||
+ | **8/12 nested-PCR product : sulAp, uvrAp | ||
+ | *Colony PCR (re) (<nowiki>#</nowiki><html><span class="part_num">13</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | *Miniprep(ligation product : <nowiki>#</nowiki><html><span class="part_num">6</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">23</span></html>) | ||
+ | *Cloning(hotstart-PCR: uvrBp , cheZ) | ||
+ | *Digest | ||
+ | **XPcut:<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">6</span>-<span class="part_num">5</span></html> | ||
+ | **EScut: sulAp, uvrAp, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">23</span></html> | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/15(Mon)=== | ||
+ | *Gel extraction(from previous digest products(8/12) : <nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span></html>, <nowiki>#</nowiki><html><span class="part_num">4</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">23</span></html>) | ||
+ | *Digest | ||
+ | **XPcut: cheZ, uvrBp(PCR products) | ||
+ | **EScut: <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> (to obtain pSB1AK3 EScut) | ||
+ | *Assay of Cell lysis construction(pLuc-Lysis device) inducted by IPTG | ||
+ | *Making Lysis buffer, Agarose gel | ||
+ | *Gel extraction(from digest products XPcut: cheZ, uvrBp(PCR products) EScut: <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> to obtain pSB1AK3 EScut) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">23</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">34</span>--</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">35</span>--</html>pSB1AK3) | ||
+ | *Transformation(ligation products:WT, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>:cheZ-, <nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span></html>) -> 37 incubation | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/16(Tue)=== | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">23</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>) | ||
+ | *Culture (<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">4</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>) | ||
+ | *Incubation (<nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">35</span>-</html>pSB1AK3) | ||
+ | *Colony PCR(additional)(previously incubated ligation products: 3, 8, 10) | ||
+ | *hotstart PCR(sulAp, uvrAp, uvrBp) | ||
+ | *Digest(SPcut: <nowiki>#</nowiki><html><span class="part_num">3</span></html>, Ecut:<nowiki>#</nowiki><html><span class="part_num">3</span></html>(control)) go to 37 incubation | ||
+ | *Ligation retry(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">29</span></html>, Negative control :<nowiki>#</nowiki><html><span class="part_num">3</span></html>) | ||
+ | *Frozen stock(<nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span></html>) | ||
+ | *Culture test(<nowiki>#</nowiki><html><span class="part_num">14</span></html> on 0.3%Agar.(km)) | ||
+ | *Ethanol precipitation: 29.9ng/ul * 1000ul | ||
+ | *Miniprep(Ligation products : <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">17</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>) | ||
+ | *Electrophoresis(Colony PCR products: 3, 8, 10) | ||
+ | *Waking from frozen stock(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, CheZ-, WT) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/17(Wed)=== | ||
+ | *Hotstart-PCR (Change conditions a little) | ||
+ | **From Genome | ||
+ | **From nested-region | ||
+ | **From parts | ||
+ | *Making Competent Cell, SOB medium | ||
+ | *Ethanol precipitation(WT: 40ng/ul) | ||
+ | *Culture from master-plate(<nowiki>#</nowiki><html><span class="part_num">6</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) -> Miniprep | ||
+ | *Assay(IPTG induced cell lysis device | ||
+ | *Gel extraction(SPcut:<nowiki>#</nowiki><html><span class="part_num">3</span></html>) | ||
+ | *Culture from Frozen stock(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Electrophresis check(Loading only DNA size markers) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">6</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/18(Thu)=== | ||
+ | *Digest(8/17<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span></html> SPcut, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html> XPcut, <nowiki>#</nowiki><html><span class="part_num">33</span></html> Ecut, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>,<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>,<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html> EScut, pSB1K3 EPcut) | ||
+ | *Hotstart-PCR (Change conditions a little) | ||
+ | **From Genome : uvrBp, cheZ | ||
+ | **From nested-region : uvrA | ||
+ | *Culture from 0809 master-plate ⑪ (<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>) → Miniprep. | ||
+ | *Making LB plate & tube with various agarose conc. (plate: 0.5, 0.7, 0.9%×2 each, tube: 0.5,1.0,1.5,2.0,3.0%×2 each)(selection: Kanamycin) | ||
+ | *Gel extraction(8/17<nowiki>#</nowiki><html><span class="part_num">3</span></html>_SP,<nowiki>#</nowiki><html><span class="part_num">33</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">33</span></html>_E, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>_ES, pSB1K3 _EP) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">1</span></html>,<nowiki>#</nowiki><html><span class="part_num">2</span></html> SPcut, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> XPcut, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> SPcut(check), EPcut(selection), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html> EScut, 0817 Gel extracted PCR product (uvrBp) EScut, 0818PCR products (uvrAp, uvrBp, cheZ) ES&XPcut) | ||
+ | *Ligation (8/17<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, 8/17<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">8</span></html>/15<nowiki>#</nowiki><html><span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, 8/15<nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">28</span>-</html>pSB1AK3) | ||
+ | *Culture for frozen stock in eppendolf tube | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/19(Fri)=== | ||
+ | *ColonyPCR(from ligation products)(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction(08/18's digest products:<nowiki>#</nowiki><html><span class="part_num">1</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">2</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP, cheZ_XP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>_EP, 8/18uvrBp_ES, 8/17uvrBp_ES, uvrAp_ES) | ||
+ | *Colony PCR retry(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3) | ||
+ | *Frozen Stock | ||
+ | *Check the list of frozen stock | ||
+ | *Assay of recAp from biobrick parts using RFP construct & transilluminator | ||
+ | *Electrophoresis(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>, <nowiki>#</nowiki><html><span class="part_num">6</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">7</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/22(Mon)=== | ||
+ | *Making SOB medium | ||
+ | *Culture for Miniprep | ||
+ | **From frozen stock(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">7</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>) | ||
+ | **From master plate(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | **From master plate retry( at 13:50 OD600=0) | ||
+ | **From ligation products (for mistake above) | ||
+ | *Miniprep, Nanodrop and Electrophoresis | ||
+ | **From frozen stock(<nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(2), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>) <nowiki>#</nowiki><html><span class="part_num">7</span></html> was a little bit weak | ||
+ | **From ligation products(<nowiki>#</nowiki><html><span class="part_num">7</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest | ||
+ | **From Frozen Stock (<nowiki>#</nowiki><html><span class="part_num">7</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">30</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>_ES) | ||
+ | **From Master plate(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3_SP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *Sulvage <nowiki>#</nowiki><html><span class="part_num">33</span></html> using protocol of transformation | ||
+ | *Culture for Frozen stock | ||
+ | **From Master plate(<nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>), 700ul, in eppendolf tube | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/23(Tue)=== | ||
+ | *Gel extraction | ||
+ | **from O.N. digest products(<nowiki>#</nowiki><html><span class="part_num">7</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">30</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3_SP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *Frozen stock | ||
+ | **From O.N. culture in eppendolf tube 700ul(<nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | **From O.N. culture(for Miniprep)(<nowiki>#</nowiki><html><span class="part_num">34</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>) | ||
+ | *Miniprep | ||
+ | **retry from master plate <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>(278.1 ng/ul), <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(188.9ng/ul), <nowiki>#</nowiki><html><span class="part_num">34</span>-</html>pSB1AK3(89.6ng/ul), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>(327.0 ng/ul) O.N. culture | ||
+ | *Digest (<nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html> XPcut, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html> Xcut, Pcut (check), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html> EScut (sulvage) ) | ||
+ | *Ligation (<nowiki>#</nowiki><html><span class="part_num">14</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">30</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | *<nowiki>#</nowiki><html><span class="part_num">33</span></html> sulvage (isolation culture) | ||
+ | *Making culture(LB amp, LB) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/24(Wed)=== | ||
+ | *Gel extraction & Electrophoresis(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_X, *PCR for amplification (or cloning) -> PCR clean-up system -> digest | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">27</span></html> (82.1 ng/ul, total = 30 ul) -> XP cut | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">28</span></html> (104.4 ng/ul, total = 30 ul) -> XP cut | ||
+ | *Making plate and amp stock ->4℃ | ||
+ | **amp 32 | ||
+ | **km 11 (km conc. = 3/4 * that it was) | ||
+ | *Miniprep and Nanodrop | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">33</span></html> 32.1 ng/ul <- there is probability that culture volume was not enough ->transfromation | ||
+ | *Luciferase assay reagents prep | ||
+ | **D-Luciferin Solution : 1ml*10 + 100ul*10 at -80℃ | ||
+ | **Coelenterazine Solution : 100ul * 10 at -20℃ | ||
+ | **Lysis buffer : 500ul * 10 at -20℃ | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">34</span></html>, <nowiki>#</nowiki><html><span class="part_num">30</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span>-</html><nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">35</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span>-</html>pSB1K3, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span></html> and Negative control) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/25(Thu)=== | ||
+ | *Renilla luciferase assay (with <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> culture) | ||
+ | *Gel extraction (<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html> XPcut) | ||
+ | *Ligation (0824 retry & <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">27</span></html>) | ||
+ | *Transformation (Ligation products into WT competent cells & <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html> miniprep product into WT cell (line-drawing spread), <nowiki>#</nowiki><html><span class="part_num">33</span></html> miniprep product into ΔcheZ cell (line-drawing spread)) | ||
+ | *Digest (<nowiki>#</nowiki><html><span class="part_num">14</span></html> XPcut) | ||
+ | *Passage(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, WT, ΔcheZ, WT in eppen, ΔcheZ in eppen) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/26(Fri)=== | ||
+ | *Renilla luciferase assay (with <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> culture) | ||
+ | *Colony PCR (<nowiki>#</nowiki><html><span class="part_num">33</span></html> Miniprep transformations) | ||
+ | *Gel extraction (<nowiki>#</nowiki><html><span class="part_num">14</span></html>_XP) | ||
+ | *Ligation (retry) (using today's competent cells, 0802 ΔcheZ cells) | ||
+ | *Cloning (cheZ) | ||
+ | *Digest (<nowiki>#</nowiki><html><span class="part_num">5</span></html> EXcut, cheZ PCR products XPcut) | ||
+ | *Passage in a shaker(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, WT, ΔcheZ) | ||
+ | *Making competent cell, LB broth | ||
+ | *Contamination check (0817 comp., 0802 comp., 0802 ⊿cheZ comp. spread on LB (amp) plates) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/27(Sat)=== | ||
+ | *Ligation & Transformation (0826 ①②③⑤⑧⑨) (into Nippon Gene comp.) | ||
+ | *Contamination check (WT FS, ΔcheZ Frozen stock) | ||
+ | *Waking from Frozen stock(WT, ΔcheZ) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/28(Sun)=== | ||
+ | *Making ΔcheZ comp., LB amp plates, TB | ||
+ | *Colony PCR & master plate preparation -> Failure!! | ||
+ | *Gel extraction (<nowiki>#</nowiki><html><span class="part_num">29</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">5</span></html>_EX) | ||
+ | *Miniprep and frozen stocks (<nowiki>#</nowiki><html><span class="part_num">33</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest (<nowiki>#</nowiki><html><span class="part_num">20</span></html> SPcut, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html> XPcut) | ||
+ | *Waking from frozen stock (WT, ⊿cheZ, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/29(Mon)=== | ||
+ | *PCR purification -> digest with XP(<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>) | ||
+ | *Ligation (higher I/V ratio, into cheZ comp.)(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>(retry), <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> to pSB1K3 (retry), <nowiki>#</nowiki><html><span class="part_num">35</span>-</html>pSB1AK3, (<nowiki>#</nowiki><html><span class="part_num">33</span></html>,<nowiki>#</nowiki><html><span class="part_num">34</span></html>)-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/30(Tue)=== | ||
+ | *Making LB amp, Aspartic acid solution (10mM), H2O2 solution (10mM) | ||
+ | *Gel extraction -> PCR was failed!(<nowiki>#</nowiki><html><span class="part_num">27</span></html>,<nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | *There was trash (from PCR) below | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> to pSB1K3, <nowiki>#</nowiki><html><span class="part_num">35</span>-</html>pSB1AK3, (<nowiki>#</nowiki><html><span class="part_num">33</span></html>,<nowiki>#</nowiki><html><span class="part_num">34</span></html>)-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *PCR -> check (5uL EP -> PCR clean-up(<nowiki>#</nowiki><html><span class="part_num">27</span></html>,<nowiki>#</nowiki><html><span class="part_num">28</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, pSB1AK3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span></html>_SP) | ||
+ | *EP for MP check(<nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">29</span></html>, pSB1AK3-<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> on Amp/Km plate, <nowiki>#</nowiki><html><span class="part_num">3</span></html> (Negative control), pSB1AK3 (Negative control) on Amp/Km plate | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html> into cheZ-) -> Line-drawing plating | ||
+ | *SOS stress (UV irradiation or H2O2)(<nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>) | ||
+ | *culture for Miniprep from master plate(<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Waking from frozen stock(<nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/08/31(Wed)=== | ||
+ | *Making LB plates(amp, km) | ||
+ | *Luciferin reagent preparation | ||
+ | *Flask A.C. for making competent cells | ||
+ | *Miniprep (<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, pSB1AK3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span></html>_SP) | ||
+ | *PCR -> clean-up -> XPcut(<nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>) | ||
+ | *Waking from frozen stock(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html> (WT), <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Plating from liquid culture(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">35</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> & Negative control) | ||
+ | *Transformation | ||
+ | **Plasmid isolation(<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(MP;1:9gradient)) | ||
+ | **Part cloning(<nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span></html>) | ||
+ | **Contamination check(pSB1AK3(EPcut), pSB1K3(EPcut)) | ||
+ | **Titer check(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(Takara comp.,1:9gradient), <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(ΔcheZ comp.0802, 1:9gradient), <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(ΔcheZ comp.0830, 1:9gradient)) | ||
+ | **Ligation products | ||
+ | <html></div></html> | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labMonth" id="labMonth_9"></html> | ||
+ | ==September== | ||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/01(Thu)=== | ||
*Colony PCR | *Colony PCR | ||
- | **<nowiki>#</nowiki> | + | **<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(MP), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">35</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span></html>(from Part cloning) |
- | + | *Gel extraction | |
- | + | **<nowiki>#</nowiki><html><span class="part_num">27</span></html>_XP ,<nowiki>#</nowiki><html><span class="part_num">28</span></html>_XP(0830); <nowiki>#</nowiki><html><span class="part_num">27</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">28</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span></html>_XP -> failed!, <nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html>_XP(0831) | |
- | + | *Making medium(SOB, LB amp plate), competent cells (WT & ΔcheZ) | |
- | **<nowiki>#</nowiki> | + | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>) |
- | + | *PCR and Column purification | |
- | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html> (Recovery from colony PCR tubes!) | |
- | + | ||
- | + | ||
- | * | + | |
- | + | ||
- | * | + | |
- | **<nowiki>#</nowiki> | + | |
- | + | ||
- | + | ||
- | + | ||
- | + | ||
*Digest | *Digest | ||
- | **<nowiki>#</nowiki> | + | **<nowiki>#</nowiki><html><span class="part_num">3</span></html> -> Failed (Mixed with DMSO!), <nowiki>#</nowiki><html><span class="part_num">20</span></html> |
- | **<nowiki>#</nowiki>20-3- | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html> |
- | * | + | *Ligation |
- | * | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>20-3- | + | *Transformation |
- | ** | + | **<nowiki>#</nowiki><html><span class="part_num">36</span></html> into Nippon Gene comp. |
- | **<nowiki>#</nowiki> | + | **Ligation products into Takara comp. (including N.C.) |
- | **<nowiki>#</nowiki>20-3- | + | **Titer check(MP product <nowiki>#</nowiki><html><span class="part_num">3</span></html> into 0901 WT comp., MP product <nowiki>#</nowiki><html><span class="part_num">20</span></html> into 0901 ΔcheZ comp.) |
+ | *Mobility assay(<nowiki>#</nowiki><html><span class="part_num">14</span>-<span class="part_num">5</span></html> (km resistance) WT/ΔcheZ cells) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/02(Fri)=== | ||
+ | *Making medium and reagent(LB 120ml(0.125, 0.25, 0.5% Agar, M9 stock(20×), M9 200ml, Vitamins Solution (100×)) | ||
+ | *Gel extraction -> Digest products were OK but extraction was failed. | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>_XP | ||
+ | *Colony PCR & Master plate(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, ΔcheZ) | ||
+ | *Hot-start PCR & Column purification -> PCR products were OK but all purified products were lost. | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html> (Recovery from colony PCR tubes) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html> (from miniprep products) | ||
+ | *PCR & Column purification | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html> (from miniprep products) | ||
+ | *Digest | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">37</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, ΔcheZ_E/P (from MP products) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span></html>_ES (from PCR products) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/03(Sat)=== | ||
*Gel extraction | *Gel extraction | ||
- | **<nowiki>#</nowiki> | + | *Ligation & Transformation |
- | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html> into Nippon Gene comp. (+ High Competent Broth 240ul) | |
- | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html> into 0902 WT comp.(+ Takara SOC broth 500ul) | |
- | + | *Assays | |
- | + | **Mobility assay (.5%, .25%, .125%) | |
- | + | **Asp assay | |
- | + | *Part Cloning (<nowiki>#</nowiki><html><span class="part_num">36</span></html>) | |
- | + | <html></div></html> | |
- | + | ||
- | + | ||
- | + | ||
- | + | ||
- | **<nowiki>#</nowiki> | + | |
- | + | ||
- | + | ||
- | **<nowiki>#</nowiki>36 | + | |
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/04(Sun)=== | ||
+ | *Hot-start PCR Pre-Mix (4×) preparation (Buffer + dNTPs + THUNDER Taq) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>) | ||
+ | *Colony Diffusion Assays | ||
+ | *GFP Live-Imaging Assays | ||
+ | *Ligation & Transformation (retry) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html> into 0901 WT comp. | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span></html> into Takara comp. | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">5</span></html> as Negative Controls into 0901 WT comp. | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">20</span></html>) | ||
+ | <html></div></html> | ||
- | =='11/9/ | + | <html><div class="labDay"></html> |
+ | ==='11/09/05(Mon)=== | ||
+ | *Colony Diffusion Assays (<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>) WT,cheZ- | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">20</span></html>) | ||
+ | *Luciferase Assays | ||
+ | **Calibrations (Firefly Luciferase standards) | ||
+ | **Background Assay | ||
+ | **Dual Luciferase Assay(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">28</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span></html>) | ||
+ | *PCR -> clean-up -> Digest(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>) | ||
+ | *IPTG-inducted Cell lysis test(IPTG+, LB plate (Positive control)) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span></html>) | ||
+ | *Making M9 -> Asp -> all failed(.125%, .25% Agar) | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/06(Tue)=== | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>) | ||
+ | *PCR (retry) (Hot-start & Normal PCR) -> Purification -> Digest(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>_ES) | ||
+ | *PCR (cloning retry)(lexA, uvrBp) | ||
+ | *Ligation & Transformation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span>-</html><nowiki>#</nowiki><html><span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span>-</html>pSB1AK3 | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">5</span></html>, pSB1AK3 (N.C.) | ||
+ | *Culture for GFP live-imaging (<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html> WT/cheZ-) -> 4℃ storage | ||
+ | *Ninhydrin sol. preparation (Not dissolvable!) -> 4℃ storage | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/07(Wed)=== | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">30</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP) | ||
+ | *Culture from master plate(<nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html> (failed)) | ||
+ | *PCR purification & Digest(lexA_XP, uvrBp_ES) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>_ES, pSB1AK3_EP, <nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP) | ||
+ | *PCR amplification -> Digest(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP (re), <nowiki>#</nowiki><html><span class="part_num">2</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">34</span></html>_SP) | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | Ligation & Transformation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">33</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">1</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">33</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span></html>, pSB1AK3 (N.C.) | ||
+ | *Ninhydrin stock solution preparation | ||
+ | **Asp conc. check (it took 5 min at 80C) | ||
+ | *Making M9 medium(.25, .125% Agar.) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/08(Thu)=== | ||
+ | *Making competent cell (WT), LB(amp) plate | ||
+ | *Frozen stocks(<nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(psB1AK3)) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">2</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction | ||
+ | **PCR product: <nowiki>#</nowiki><html><span class="part_num">35</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>_XP | ||
+ | **Miniprep product: <nowiki>#</nowiki><html><span class="part_num">2</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">34</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES | ||
+ | *PCR amplification(<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>) | ||
+ | *PCR for Sequencing -> Store at -20C(<nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">35</span></html>_ES-psB1AK3_ES, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_XP-psB1A2_XP, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_XP-psB1AK3_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest | ||
+ | **PCR products: <nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES | ||
+ | **pSB1C3_EP | ||
+ | *Asp Gradient test | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/09(Fri)=== | ||
+ | *Making competent cell (WT), TB | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">35</span>-</html>pSB1AK3 (failed), <nowiki>#</nowiki><html><span class="part_num">31</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-</html><nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>(failed)) | ||
+ | *PCR amplification | ||
+ | **From Miniprep : <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | **From Colony PCR : <nowiki>#</nowiki><html><span class="part_num">31</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (failed) | ||
+ | *Digest | ||
+ | **Miniprep product: <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>_EP | ||
+ | **PCR-purified product: <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> (pSB1AK3)) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>_EP) | ||
+ | *Ligation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">29</span></html>_XP-pSB1AK3_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP(retry) | ||
+ | *Comp. check | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> (pSB1AK3) into 0908 WT comp. | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> into 0909 WT comp. | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> into 0802 cheZ- comp. | ||
+ | *Part cloning(BBa_J04450 (IPTG-induced RFP reporter on pSB1C3)) | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">31</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *UV irradiation -> Luciferase Assay(<nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Asp Gradient test | ||
+ | *CleanSEQ -> Sequencing(<nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">34</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">37</span>-<span class="part_num">5</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/10(Sat)=== | ||
+ | *Culture plate storage (4℃) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">31</span>-</html>pSB1AK3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (Maybe Ligation was FAILURE , estimated length is 1000-1500 by electrophoresis)) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/11(Sun)=== | ||
+ | *Digest from Miniprep products | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP -> O.N. | ||
*Colony PCR | *Colony PCR | ||
- | **<nowiki>#</nowiki>36-3-14-5-2-3-17- | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> (pSB1C3), <nowiki>#</nowiki><html><span class="part_num">29</span></html> (pSB1AK3), BBa_J04450 (pSB1C3) |
- | **<nowiki>#</nowiki>36-3-14-5-2-3-17- | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>(failed), <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>3-27- | + | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP) |
- | **<nowiki>#</nowiki>36-3-29- | + | *Culture for Miniprep |
+ | **From Master plate : <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">29</span></html> (pSB1AK3), BBa_J04450 (pSB1C3) | ||
+ | **From plate stock : <nowiki>#</nowiki><html><span class="part_num">20</span></html> | ||
+ | **From frozen stock : <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | *Miniprep( <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">1</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *Direct Ligation | ||
+ | **i.PCR amplification -> Digest for >1h -> Heat kill for 20min (<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES) | ||
+ | **ii.Digest for >1h -> Heat kill for 20min (<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_SP, pSB1A3_EP, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX) | ||
+ | **iii.Ligation for 30min (<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1A3_EP) | ||
+ | **iv.Transformation | ||
+ | *Making 0.125% agar LB amp plate (IPTG 1, 10, 30, 100uM) | ||
+ | *Cell Diffusion Assay | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> WT (IPTG 1, 10, 30, 100uM) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> cheZ-/- (IPTG 1, 10, 30, 100uM) | ||
+ | **WT | ||
+ | **cheZ-/- | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/9/12 (Mon)=== | ||
+ | *Colony PCR | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">1</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> (no colony) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span>-</html>pSB1A3 (PCR failed?) | ||
+ | Note: <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> colonies on the master-plate are RED. Maybe <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>...<br> | ||
+ | Note: <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span>-</html>pSB1A3 PCR amplification doesn't work. Seems failed extension... | ||
+ | *PCR amplification | ||
+ | **From master plate : <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> | ||
+ | **From 09/11 Miniprep product : <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | Note: Only one <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> colony is NOT RED. | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">29</span></html> (pSB1AK3), <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, BBa_J04450 (pSB1C3)) | ||
+ | *Digest | ||
+ | **From PCR : <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP | ||
+ | **pSB1A3_EP, pSB1C3_EP | ||
+ | **From Miniprep : <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP | ||
+ | **J04450_EP, *<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES | ||
+ | *Gel extraction | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_SP | ||
+ | **From OverNight digests : <nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES | ||
+ | **From today's PCR digests : <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> from 0911 master plate) | ||
+ | *Dual Luciferase Assay(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span>-</html>pSB1A3) | ||
+ | *Ligation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1A3_EP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_EP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP-<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1AK3_EP | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/9/13 (Tue)=== | ||
+ | *Colony PCR | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1A3_EP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_EP (failed) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP-<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP (failed) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1AK3_EP | ||
*Culture for Luciferase Assay | *Culture for Luciferase Assay | ||
- | **<nowiki>#</nowiki>36-3-14-5-2-3-17-5-20-3-37-5(pSB1A3, pSB1C3) (IPTG 0, 1, 10, 38, 100uM) | + | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>(pSB1A3, pSB1C3) (IPTG 0, 1, 10, 38, 100uM) |
- | **<nowiki>#</nowiki>20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM) | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0, 1, 10, 38, 100uM) |
- | **<nowiki>#</nowiki>1-3-14-5-2-3-17-5 (IPTG 0uM)<br> | + | **<nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0uM)<br> |
Note: Seemes OD600 should NOT exceed 0.4~0.5, or intracellular luciferase will be saturated. | Note: Seemes OD600 should NOT exceed 0.4~0.5, or intracellular luciferase will be saturated. | ||
*Miniprep. | *Miniprep. | ||
- | **<nowiki>#</nowiki>20-3-14-5-2-3-17-5 | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>1-3-14-5-2-3-17-5 | + | **<nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>20-3-37-5 (from a non-red colony on the master plate) | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> (from a non-red colony on the master plate) |
- | *Gel | + | *Gel extraction |
- | **<nowiki>#</nowiki>20-3-37- | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP (from non-red colony) |
- | **<nowiki>#</nowiki>20-3-29- | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES(failed) |
- | **<nowiki>#</nowiki>2- | + | **<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">9</span></html>_XP, pSB1C3_EP |
- | Note: <nowiki>#</nowiki>20-3-29-5 was not cut! | + | Note: <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> was not cut! |
*Dual Luciferase Assay | *Dual Luciferase Assay | ||
- | **<nowiki>#</nowiki>1-3-14-5-2-3-17-5 (IPTG 0uM) | + | **<nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0uM) |
- | **<nowiki>#</nowiki>20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM) | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0, 1, 10, 38, 100uM) |
- | **<nowiki>#</nowiki>36-3-14-5-2-3-17-5-20-3-37-5 (<nowiki>#</nowiki>2-3-11-5?) (IPTG 0, 1, 10, 38, 100uM) | + | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> (<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>?) (IPTG 0, 1, 10, 38, 100uM) |
*Sequencing preparation | *Sequencing preparation | ||
- | **<nowiki>#</nowiki>20-3-37-5 (from a non-red colony on the master plate) | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> (from a non-red colony on the master plate) |
- | **<nowiki>#</nowiki>20-3-29-5 | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>3-27-5 | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>31 | + | **<nowiki>#</nowiki><html><span class="part_num">31</span></html> |
*Digest | *Digest | ||
- | **<nowiki>#</nowiki> | + | **<nowiki>#</nowiki><html><span class="part_num">20</span></html>_E |
- | **<nowiki>#</nowiki>3-29- | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_S (from MP products) (cut check) |
*Ligation | *Ligation | ||
- | **<nowiki>#</nowiki> | + | **<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP |
- | *Transformation -> | + | *Transformation |
- | **<nowiki>#</nowiki>3-14-5-2-3-17-5 | + | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>1-3-14-5-2-3-17-5 | + | **<nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> |
- | **<nowiki>#</nowiki>20-3-14-5-2-3-17-5 (for plate stock) | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (for plate stock) |
- | **<nowiki>#</nowiki>20-3-37-5 (from a non-red colony on the master plate) (for | + | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> (from a non-red colony on the master plate) (for plasmid check) |
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/14(Wed)=== | ||
+ | *Colony PCR | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP-<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP (failed) | ||
+ | *Culture for Luciferase Assay (from plate stocks) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0, 1, 10, 38, 100uM) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0uM) | ||
+ | *Digest | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (EPcut) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP | ||
+ | *Culture for Miniprep. | ||
+ | **From plate stock : <nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> | ||
+ | **From 09/11 master plate : <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> (pSB1C3) | ||
+ | *Part cloning(<nowiki>#</nowiki><html><span class="part_num">19</span></html>: BBa_R0010) | ||
+ | *Ligation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3 | ||
+ | *Making WT competent cell | ||
+ | *Dual Luciferase Assay | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0uM) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (IPTG 0, 1, 10, 38, 100uM) | ||
+ | *Asp TLC Assay | ||
+ | *Asp Diffusion Assay (0.25% Agar) | ||
+ | *Fumaric Acid (Fum) Sol. preparation | ||
+ | *Sequencing | ||
+ | **①F⑤R<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> (from a non-red colony on the master plate) | ||
+ | **②F⑥R<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> (<nowiki>#</nowiki><html><span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>?) | ||
+ | **③F⑦R<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html> | ||
+ | **④F⑧R<nowiki>#</nowiki><html><span class="part_num">31</span></html> | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/15(Thu)=== | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">19</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> (failed), <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">20</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>(failed), <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (EPcut); <nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP | ||
+ | *Making WT competent cell | ||
+ | *Making LB plates(amp:39, cm:18) | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> (pSB1C3), <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">19</span></html>) | ||
+ | *Ligation | ||
+ | **sulAp(ES)-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP-pSB1C3_PCR digest(EP) | ||
+ | **recNp(ES)-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP-pSB1C3_PCR digest(EP) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_PCR digest(EP) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP-<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP) | ||
+ | *Culture for Dual Luciferase Assay(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *TLC check (LB, M9) | ||
+ | *Cell Diffusion Assay | ||
+ | *Racing Assay (0.25% Agar, 10mM Asp Sol.) | ||
+ | *Fumaric Acid (Fum) Sol. preparation (failed!!) | ||
+ | *Sequencing | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/16(Fri)=== | ||
+ | *Colony PCR | ||
+ | **sulAp(ES)-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP-pSB1C3_PCR digest(EP), | ||
+ | **recNp(ES)-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP-pSB1C3_PCR digest(EP), | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_PCR digest(EP), | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP-<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP), | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP), | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP), | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_PCR digest(EP) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_PCR digest(EP) (retry; non-red colonies) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html> (pSB1C3), <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">19</span></html>) | ||
+ | *PCR amplification(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, pSB1C3) | ||
+ | *Sequencing prep(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>) | ||
+ | *Making agarose gel, WT competent cell, | ||
+ | *Electrophoresis check(pSB1C3_PCR) | ||
+ | *Digest(pSB1C3_ES, pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES) | ||
+ | *Dual luciferase assay | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (1h, 1.5h culture)(IPTG 0, 1, 10, 100uM)(2.5mL) | ||
+ | *Fumaric Acid (Fum) Sol. preparation | ||
+ | **TLC test | ||
+ | **Asp diffusion test | ||
+ | **Asp racing assay | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/17(Sat)=== | ||
+ | *Colony PCR (retry) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (pSB1C3) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> | ||
+ | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (pSB1C3)) | ||
+ | *Ligation(sulAp-pSB1C3, recNp-pSB1C3, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> into cheZ-) | ||
+ | *Making medium(.25% Agar. M9& LB each 20 plates) | ||
+ | *Cell diffusion assay | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/18(Sun)=== | ||
+ | *Colony PCR & Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> (pSB1C3)) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, recNp-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, sulAp-pSB1C3, recNp-pSB1C3, <nowiki>#</nowiki><html><span class="part_num">31</span>-</html>pSB1C3) | ||
+ | *Cell diffusion assay | ||
+ | **Innoculate colony(WT / cheZ-/- / cheZ-/- + cheZ plasmid) | ||
+ | *GFP imaging assay(Culture cells expressing GFP) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/19(Mon)=== | ||
+ | *Culture for Dual luciferase assay(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">1</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | *Sequencing(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>) | ||
+ | *Dual luciferase assay(<nowiki>#</nowiki><html><span class="part_num">1</span></html>~, <nowiki>#</nowiki><html><span class="part_num">20</span></html>~, <nowiki>#</nowiki><html><span class="part_num">36</span></html>~, <nowiki>#</nowiki><html><span class="part_num">33</span></html>~ ) | ||
+ | *Asp racing assay(Innoculate colonies) | ||
+ | *GFP imaging assay<html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/20(Tue)=== | ||
+ | *colony PCR(20-3-27-5, <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *PCR amplification -> Digest(<nowiki>#</nowiki><html><span class="part_num">29</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP) | ||
+ | *Sequencing prep. | ||
+ | *Ethanol precipitation (10ul TE) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>(pSB1C3): 144.5ng/uL | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>: 728.0ng/uL | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(pSB1C3): 505.7ng/uL | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">31</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_EP) | ||
+ | *Luciferase assay | ||
+ | *Asp diffusion assay | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/21(Wed)=== | ||
+ | *Making plates | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">29</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP) | ||
+ | *Ligation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">24</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP, <nowiki>#</nowiki><html><span class="part_num">29</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP | ||
+ | *Transformation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span></html> from BioBrick? part | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html> into BL21 comp. | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES) | ||
+ | *Luciferase Assay(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Cell diffusion assay | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/22(Thu)=== | ||
+ | *Ligation (retry) | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP-<nowiki>#</nowiki><html><span class="part_num">20</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-</html><nowiki>#</nowiki><html><span class="part_num">24</span></html>_SP, <nowiki>#</nowiki><html><span class="part_num">29</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">36</span></html>(->culture for mini prep), <nowiki>#</nowiki><html><span class="part_num">33</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>(->Maybe the amplification was failed)) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_SP, _ES; <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES) | ||
+ | *Making plates | ||
+ | **M9 0.25% Agar. +IPTG +Amp.: 20 | ||
+ | **LB Cm.: 20 | ||
+ | *Ligation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP, | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3, | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_EP | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/23(Fri)=== | ||
+ | *colony PCR | ||
+ | *Column purification | ||
+ | *Ligation | ||
+ | *PCR amplification | ||
+ | *Competent cell(BL21(DE3)) | ||
+ | *Asp diffusion test | ||
+ | *Cell diffusion assay | ||
+ | *Asp chemotaxis assay | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/26(Mon)=== | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">20</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-</html><nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, recNp-3-14-5-2-3-17-5) | ||
+ | *Digest(pSB1C3_ES) | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span>-</html>pSB1C3, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span>-</html>pSB1C3, <nowiki>#</nowiki><html><span class="part_num">36</span></html>) | ||
+ | *PCR amplification(pSB1A3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>) | ||
+ | *Ethanol precipitation(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span>-</html>pSB1C3, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span>-</html>pSB1C3) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/27(Tue)=== | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">35</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP, pSB1C3_ES, pSB1C3_XP) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_SP, pSB1C3_ES, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_EP-pSB1C3_EP) | ||
+ | *Gel digestion(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP, pSB1C3_ES, pSB1C3_XP) | ||
+ | *Nanodrop(<nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP, pSB1C3) | ||
+ | *PCR amplification(J04450) | ||
+ | *Gel extraction(pSB1C3_ES) | ||
+ | |||
+ | <html></div></html> | ||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/28(Wed)=== | ||
+ | *Nanofrop | ||
+ | **From PCR : <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, pSB1C3 | ||
+ | **From Miniprep : <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, J04450 | ||
+ | *Transformation(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">5</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) -> BL21(DE) comp. | ||
+ | *PCR amplification and Digest(EPcut) (<nowiki>#</nowiki><html><span class="part_num">31</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest(pSB1C3_EP, pSB1C3_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">24</span></html>_SP_SP, <nowiki>#</nowiki><html><span class="part_num">24</span></html>_SP) | ||
+ | *Sequencing(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Making AspA culture | ||
+ | *Cell diffusion assay | ||
+ | *Asp taxis assay | ||
+ | *Gel extraction and Nanodrop(pSB1C3_EP, pSB1C3_XP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES, <nowiki>#</nowiki><html><span class="part_num">24</span></html>_SP) | ||
+ | *Ligation | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_EP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP-pSB1C3_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_XP-pSB1C3_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">29</span></html>_XP-pSB1C3_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">24</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP | ||
+ | **<nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_XP | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/29(Thu)=== | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP) | ||
+ | *Gel extraction and Nanodrop(<nowiki>#</nowiki><html><span class="part_num">31</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_EP) | ||
+ | *PCR amplification(pSB1C3, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Digest(pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *Colony PCR, PCR amplification, Miniprep and Colony diffusion assay(<nowiki>#</nowiki><html><span class="part_num">36</span></html>_SP-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_XP) | ||
+ | *Colony PCR and Miniprep(<nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/09/30(Fri)=== | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES) | ||
+ | *PCR amplification and Digest(EPcut)(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">31</span></html>) | ||
+ | *Digest(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES, pSB1C3_EP, pSB1C3_ES) | ||
+ | *Culture for MIniprep from plate stock(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Making competent cell(JM109) | ||
+ | *Gel extraction and Nanodrop(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES_PCR, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES_Miniprep, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP, pSB1C3_EP, pSB1C3_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">29</span></html>_EP, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_EP) | ||
+ | *Ligation(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">29</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">31</span></html>_EP-pSB1C3_EP, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>_ES-<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>_XP-pSB1C3_EP, recNp_ES-pSB1C3_ES, sulAp_ES-pSB1C3_ES, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP) | ||
+ | *Sequencing prepartion(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_F, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> | ||
+ | *Miniprep, Digest(EXcut) and Nanodrop(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Culture for Miniprep(pSB1C3, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>) | ||
+ | *Making plate LB Cm | ||
+ | <html></div></html> | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labMonth" id="labMonth_10"></html> | ||
+ | ==October== | ||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/10/01(Sat)=== | ||
+ | *Part Sequencing for parts shipping | ||
+ | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>) | ||
+ | *Sequencing preparation and Colony PCR(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>_F, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">24</span>-<span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *second colony PCR | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/10/02(Sun)=== | ||
+ | *PCR amplification(using Quick Taq), Digest(Epcut), Gel extraction and Nanodrop(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>) | ||
+ | *Gel extraction(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX) | ||
+ | *Dual Luciferase assay(<nowiki>#</nowiki><html><span class="part_num">24</span></html>) | ||
+ | *<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html> Assay | ||
+ | *<nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html> Assay | ||
+ | *Ligation(recNp_ES-<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EX, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>_EP-pSB1C3_EP) | ||
+ | *Making agarose gel | ||
+ | *Asp degradation assay | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/10/03(Mon)=== | ||
+ | *Miniprep, Nanodrop and PCR sequencing(<nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">31</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, recNp, sulAp) | ||
+ | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span>-</html>pSB1C3) | ||
+ | *Preparation for shipping | ||
+ | <html></div></html> | ||
+ | |||
+ | <html><div class="labDay"></html> | ||
+ | ==='11/10/04(Tue)=== | ||
+ | *Preparation for shipping(<nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">11</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">14</span>-<span class="part_num">5</span>-<span class="part_num">2</span>-<span class="part_num">3</span>-<span class="part_num">17</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">29</span></html>, <nowiki>#</nowiki><html><span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">36</span>-<span class="part_num">3</span>-<span class="part_num">29</span>-<span class="part_num">5</span>-<span class="part_num">20</span>-<span class="part_num">3</span>-<span class="part_num">37</span>-<span class="part_num">5</span></html>, <nowiki>#</nowiki><html><span class="part_num">31</span></html>, <nowiki>#</nowiki><html><span class="part_num">33</span></html>, <nowiki>#</nowiki><html><span class="part_num">35</span></html>) | ||
+ | <html></div></html> | ||
+ | |||
+ | <html></div></html> <!-- end of div#labMonth --> | ||
<html> | <html> | ||
Line 250: | Line 1,310: | ||
color: #fff; | color: #fff; | ||
} | } | ||
+ | |||
+ | #calendar table { | ||
+ | font: 16px/20px "Segoe UI",Calibri,"Myriad Pro",Myriad,Frutiger,"Frutiger Linotype","Trebuchet MS",Helvetica,Arial,sans-serif | ||
+ | } | ||
+ | |||
+ | #calendar table tr td.other-month { | ||
+ | color: #888; | ||
+ | } | ||
+ | |||
+ | #calendar table tr td { | ||
+ | padding: 10px; | ||
+ | } | ||
+ | |||
+ | #calendar span.clickable { | ||
+ | cursor: pointer; | ||
+ | color: #7FB9FF; | ||
+ | } | ||
+ | |||
+ | ul#months li { | ||
+ | list-style: none; | ||
+ | float: left; | ||
+ | font-size: 18px; | ||
+ | color: #7FB9FF; | ||
+ | margin-right:20px; | ||
+ | } | ||
+ | |||
+ | ul#months li.active { | ||
+ | font-weight: bold; | ||
+ | color: #fff; | ||
+ | } | ||
+ | |||
</style> | </style> | ||
<script type="text/javascript" src="http://igem-ut.net/2011/jquery.qtip-1.0.0-rc3.min.js"></script> | <script type="text/javascript" src="http://igem-ut.net/2011/jquery.qtip-1.0.0-rc3.min.js"></script> | ||
+ | <script type="text/javascript" src="http://igem-ut.net/2011/jquery.calendar-widget.js"></script> | ||
+ | |||
<script type="text/javascript"> | <script type="text/javascript"> | ||
$(function() { | $(function() { | ||
- | init(); | + | init(); |
}); | }); | ||
function init() { | function init() { | ||
- | $(' | + | // calendar |
- | + | $('ul#months li').each(function(i) { | |
- | + | $(this).css('cursor','pointer').click(function() { | |
- | + | $('#calendar').calendarWidget({month: i+4, yaer: 2011}); | |
- | + | $('ul#months li').removeClass('active'); | |
- | + | $('.labMonth').hide(); | |
- | + | $('#labMonth_'+(i+5)).show(); | |
- | + | $('#labMonth_'+(i+5)+' .labDay h3').each(function() { | |
- | + | var m = $(this).text().match(/\/(\d+) *\(/); | |
- | + | if (!m) return; | |
- | + | var tag = m[1]; | |
- | + | var target = this; | |
+ | tag = tag.replace(/^0+/,''); | ||
+ | $('#calendar td.day'+tag+' span.day').addClass('clickable').click(function() { | ||
+ | $.scrollTo(target, 500); | ||
+ | }); | ||
+ | }); | ||
+ | $(this).addClass('active'); | ||
+ | }); | ||
}); | }); | ||
+ | $('ul#months li').first().click(); | ||
+ | |||
+ | // part description popup | ||
var description = {}; | var description = {}; | ||
$('#partslist tr td:first-child').each(function() { | $('#partslist tr td:first-child').each(function() { | ||
Line 277: | Line 1,380: | ||
var num = m[1]; | var num = m[1]; | ||
var msg = ''; | var msg = ''; | ||
- | var idx2name = { 0: ' | + | var idx2name = { 0: 'Registry ID', 1: 'Content', 2: 'Plasmid', 3: 'Length' }; |
$(this).siblings().each(function(i) { | $(this).siblings().each(function(i) { | ||
msg += idx2name[i] + ': ' + $(this).html() + '<br />'; | msg += idx2name[i] + ': ' + $(this).html() + '<br />'; | ||
}); | }); | ||
- | |||
description[num] = msg; | description[num] = msg; | ||
}); | }); | ||
Line 288: | Line 1,390: | ||
}); | }); | ||
} | } | ||
- | </script> | + | </script></html> |
- | </html> | + | |
{{:Team:UT-Tokyo/Templates/EndContent}} | {{:Team:UT-Tokyo/Templates/EndContent}} |
Latest revision as of 20:25, 5 October 2011
Lab Note
![logo](https://static.igem.org/mediawiki/2011/5/5c/UT-Tokyo_logo.png)
iGEM UT-Tokyo
Parts List
Each part is represented by s number (e.g. #20 for lac promotor) in this notebook. By pointing the mouse on the number within notebook entires, you can find out the details of the part.
Number | Reg.ID | Content | Plasmid | Length (bp) |
---|---|---|---|---|
#1 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23119 J23119] | c.Promoter (Strong) | pSB1A2 | 35 |
#2 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_J23118 J23118] | c.Promoter (Medium) | [http://partsregistry.org/wiki/index.php?title=Part:BBa_J61002 BBa_J61002] | 35 |
#3 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_B0032 B0032] | RBS | pSB1A2 | 13 |
#5 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_B0014 B0014] | d.Ter | pSB1AK3 | 95 |
#9 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_E0240 E0240] | RBS-GFP-d.Ter | pSB1A2 | 876 |
#10 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_E0040 E0040] | GFP | pSB1A2 | 681 |
#11 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_E1010 E1010] | RFP | pSB2K3 | 723 |
#14 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_I712019 I712019] | fLuc | pSB1AK8 | 1653 |
#17 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_J52008 J52008] | rLuc | pSB1AK3 | 936 |
#20 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0011 R0011] | lacP | pSB1A2 | 55 |
#21 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0012 C0012] | LacI | pSB1A2 | 1128 |
#22 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_I712074 I712074] | pT7 | pSB1AK8 | 46 |
#23 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_K145001 K145001] | T7 RNA Pol. | pSB1A2 | 2655 |
#24 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_J22106 J22106] | recAp | pSB1A2 | 192 |
#27 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0083 C0083] | AspA | pSB2K3 | 1518 |
#28 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_K112808 K112808] | T4 phage lysis device (no promoter) | pSB1A2 | 1785 |
#29 | - | CheZ | pSB1AK3 | 728 |
#30 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_K117000 K117000] | Lysis gene | pSB1A2 | 144 |
#31 | - | LexA | pSB1AK3 | 750 |
#33 | - | sulAp | pSB1AK3 | 67 |
#34 | - | uvrAp | pSB1AK3 | 96 |
#35 | - | recNp | ||
#36 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0051 R0051] | cI-repressed promoter | pSB1A2 | 49 |
#37 | [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0051 C0051] | cI repressor (LVA tagged) | pSB1A2 | \ |
#38 | - | RecA |
Lab Notebook
- May
- June
- July
- August
- September
- October
Please enable Javascript to view this calendar.
May
'11/05/18 (Wed)
- Making LB medium, 50×TAE, Tris-HCl (pH8.0)
'11/05/24 (Tue)
- Making SOB medium, 0.5M EDTA (pH8.0)
'11/05/25(Wed)
- Making TB(pH=6.7), LB plate
'11/05/26(Thu)
- Making Mgaq(for SOB medium), Competent cell
- TB filtration
'11/05/31(Tue)
- iGEM parts resuspension + frozen stock (at -20℃)
- Transformation
- Overnight culture on LB plate (with 100ug/ml ampicilline)
June
'11/06/01(Wed)
- Picking up colony and transfer to LB medium
- Making LB medium
'11/06/02(Thu)
- Miniprep
- Making Glycerol(50%)(for cryopreservation)
'11/06/07(Tue)
- Nanodrop
- Transformation
- Culture from frozen stock
'11/06/08(Wed)
- Picking up colony
'11/06/09(Thu)
- Miniprep
'11/06/13(Mon)
- Planting Negative control
- Making frozen stock
- Transformation(BBa_E0240, #3, B0015, #5, #10, ,J31000, ,J44000)
'11/06/14(Tue)
- Picking up colony
- Making Mg reagent
'11/06/15(Wed)
- Miniprep
'11/06/17(Fri)
- Picking up colony(#3, B0015, B0040,#9, J31000, J44000)
- Miniprep(#5)
- Dissolution(#1, #2, K16500, #24)
'11/06/21(Tue)
- Making frozen stock(#3, B0015, B0040, #9, J31000, J44000)
- Passafe culture(#9, J31000)
- Nanodrop again
'11/06/22(Wed)
- Miniprep(#9, J31000)
- Making competent cell
'11/06/24(Fri)
- Digest (product of Miniprep 06/09 EScut)
- Making agarose gel
- Electrophoresis
'11/06/28(Tue)
- Digest(product of Miniprep 06/09 EPcut)
- Electrophoresis
- Defrost and transformation(BBa_E0030, E0020, #14, I712052,#17)
- Making 1×TAE, agarose gel, LB medium, LB plate(amp),
'11/06/29(Wed)
- Picking up colony
- Making LB broth
July
'11/07/01(Fri)
- Digest(EPcut)
- Miniprep(BBa_E0030, E0020, #14, I712052, #17)
'11/07/05(Tue)
- Electrophoresis(product of digest 07/01)
- Digest(#9, #14, #17, #3, #5)
- Making antibiotic stock(1000×)(Km,Cm)
'11/07/06(Wed)
- Gel extraction (pre)(BBa_I712052)
- Picking up colony(Ba_J23119, #2, #24, #10)
- Making agarose gel
'11/07/07(Thu)
- Digest(BBa_E0240, #14, #17, #3, #5)
- Defrost(BBa_E1010, K325101, #20, #21, #22, #23)
- Transformation(BBa_K145001, #20, #21, #22)
- Making LB plate(Cm,Km)
'11/07/08(Fri)
- Miniprep(BBa_J23119, #2, #10, #24)
'11/07/11(Mon)
- Gel extraction(#9 #14, #17)
- Transformation(#1, #11, #20, #21, K325101, #22, #23)
'11/07/12(Tue)
- Picking up colony
'11/07/13(Wed)
- Frozen stock(#1(18A), #20(6G), #21(2O), #14(6N), #23(2F))
- Picking up colony(#11)
- Gel extraction(B#3, #5)
- Digest (#2 SPcut)
'11/07/14(Thu)
- Miniprep(#1, #11, #20, #21, #22, #23)
- Electrophoresis(#2)
- Gel extraction(#2, #3, #5)
- Passafe(#11)
- Making LB+amp
'11/07/15(Fri)
- Ligation(#2-#9, #2-#3, #14-#5, #17-#5)
- Transformation(#2-#9, #2-#3, #14-#5, #17-#5)
- Dispensing Ligation Buffer
- Frozen stock(#11)
'11/07/19(Tue)
- Digest
- Gel extraction
- Making competent cell
'11/07/20(Wed)
- Picking up colony
- Making Master plate
- Testing product of Miniprep
- Transformation (#2, #27, #28)
'11/07/21(Thu)
- Miniprep(#2-3, #2-9, #14-5, #17-5, #11, #20)
- Testing product
- Digest
- SPcut : #20, #2-3
- XPcut : #10, #11, #14-5, #17-5
- Making TB
'11/07/22(Fri)
- Frozen stock(#2, #2-3, #2-9, #14-5, #17-5)
- Electrophoresis(#20_SP, #2-3_SP, #10_XP, #11_XP, #14-5_XP, #17-5_XP)
- Digest
- SPcut : #20
- XPcut : #10, #11, #14-5, #17-5
- Transformation(#27, #28, product of Miniprep 07/20)
'11/07/25(Mon)
- Colony check(#2, #2-9)
- Gel Extraction(#20_SP, #10_XP, #11_XP, #14-5_XP, #17-5_XP)
- Ligation and Transformation(#20-#9, #3-#11, #3-#14-5, #3-#17-5)
- Defrost Primer(200×, 10×)
- Dispensing PCR Mix
'11/07/26(Tue)
- Picking up colony and making master plate(#20-9, #3-11, #3-14-5, #3-17-5)
- Colony PCR(#20-9, #3-11, #3-14-5, #3-17-5)
- Digest(#14-5 XPcut,Ecut, #22 SPcut, #23 XPcut)
- Ligation(#3-#14-5(Re), #3(Negative control))
- Transformation(#15, #27, #28, #30, #3-14-5(Re), #3(Negative control))
- Making reagent for TB(KOH, CaCl2, KCl, MnCl2)
'11/07/27(Wed)
- Miniprep(#1, #2, #3, #10, #22, #24, #20-9, #3-11, #3-17-5)
- Electrophoresis
'11/07/28(Thu)
- Gel extraction(#14-5_XP, #22_SP, #23_XP)
- Digest
- SPcut : #1,#2, #3, #24
- EScut : #3-11
- XPcut : #3-17-5
- Ligation(#3-#14-5)
- Making TB bugger, LB plate, 0.3% LB plate
'11/07/29(Fri)
- Cloning(lexA, cheZ)
- Colony PCR(#3-#14-5)
- Gel extraction(#1_SP, #2_SP, #3_SP, #24_SP, #3-11_ES, #3-17-5_XP, lexA, cheZ PCR product)
- Miniprep(#30)
- Digest(lexA, cheZ PCR product XP cut)
August
'11/08/01(Mon)
- Nanodrop(#3-11(1), #1, #3, #24, #2, #3-17-5(4)M#30)
- Digest(#3 SPcut, #5 EXcut, #9 XPcut, #30 XP cut)
- Cloning(lexA, cheZ)
- Gel extraction(#3, #9, #30, pSB1A2)
- Ligation(#2-#3-17-5, #3-11-#5, #14-#5, #3-#23, #22-#9, #3-#30)
- Transformation(#2-#3-17-5, #3-11-#5, #14-#5, #3-#23, #22-#9, #3-#30, #27(IPTG), #28)
'11/08/02(Tue)
- Colony PCR(#2-#3-17-5, #3-11-#5, #14-#5, #3-#23, #22-#9)
- Cloning(PCR)(#27, #28)
- Gel extraction
- PCR_2nd(lexA, cheZ, #27, #28)
- Digest (#30 again)
- Making competent cell(cheZ-, Wild Type)
- Miniprep
'11/08/04(Thu)
- Electrophoresis(product of ligation #2-3-17-5(2), #3-11-5(1,2), #14-5(2), #22-9(1,2,3))
- Frozen stock(#2-3-17-5(2), #3-11-5(1,2), #14-5(2), #22-9(1,2))
- Making Gel
- Digest(#23 XPcut)
- IPTG induct(#20-9)
'11/08/05(Fri)
- Digest(#14-5(2) XPcut, #3-11-5(1) XPcut)
- Cloning(PCR)(#27, #28, lexA, cheZ)
'11/08/08(Mon)
- Defrost Frozen stock, Miniprep, Nanodrop and Electrophoresis(#5, #20, #23, #30)
- Making Gel
- Digest
- XPcut : #27, #28, cheZ, lexA, #3-11-5(1), #23,#30
- EScut : #6
- EXcut : #7
- SPcut : #7,#20
- Defrost Frozen stock(#5, #14-5(2), #3-11-5(1))
'11/08/09(Tue)
- Miniprep, Electrophoresis and Digest(#5, #14-5(2), #3-11-5(1))
- Gel extraction and Nanodrop(#6, #27, #8, cheZ, lexA③, #3-11-5(1), #23, #7_EX, #20, #7_SP)
- Ligation(#27-psB1A2, #3-27, #20-28, #28-psB1A2, #3-29, #29-psB1A2, #31-psB1A2, #3-#23, #1-#3-11-5(1), #2-#3-11-5(1), #24-#3-11-5(1), #3-#30)
- Making LB plate
'11/08/10(Wed)
- Colony PCR(#27-pSB1A2, #3-27, #20-28, #28-psB1A2, #3-29, #29-psB1A2, #31-pSB1A2, #3-#23, #1-#3-11-5(1), #2-#3-11-5(1), #24-#3-11-5(1), #3-#30))
- Cloning(lexA, cheZ)
- Digest
- EScut : #6
- EXcut : #7
- SPcut : #1, #2, #3, #7, #20
- XPcut : #23
- Defrost Primer
'11/08/11(Thu)
- Gel extraction(#30, #14-5(insert,vector), #5, #3-11-5, cheZ, lexA)
- Cloning(sulAp, uvrAp)
- Miniprep(#3, #20-28, #31, #24-3-11-5, #3-30)
'11/08/12(Fri)
- Colony PCR(#3-14-5(2), #6-5, #3-23)
- Cloning(nested-PCR)(uvrAp, sulAp)
- Check Electrophoresis
- 8/11 PCR product:#27, #28
- Miniprep product:#3-14-5(2), #5
- Digest(#5 EXcut, #14 EScut)
- Gel extraction
- 8/11 digest product : #5, #14-5, #3-30, cheZ, lexA
- 8/11 PCR product : #27, #28
- 8/12 nested-PCR product : sulAp, uvrAp
- Colony PCR (re) (#13-14-5)
- Miniprep(ligation product : #6-5, #3-23)
- Cloning(hotstart-PCR: uvrBp , cheZ)
- Digest
- XPcut:#27, #28, #6-5
- EScut: sulAp, uvrAp, #3-23
'11/08/15(Mon)
- Gel extraction(from previous digest products(8/12) : #27, #28, #33, #34, #4, #3-23)
- Digest
- XPcut: cheZ, uvrBp(PCR products)
- EScut: #3-11-5 (to obtain pSB1AK3 EScut)
- Assay of Cell lysis construction(pLuc-Lysis device) inducted by IPTG
- Making Lysis buffer, Agarose gel
- Gel extraction(from digest products XPcut: cheZ, uvrBp(PCR products) EScut: #3-11-5 to obtain pSB1AK3 EScut)
- Ligation(#3-#14-5, #24-#3-17-5, #24-#3-11-5, #20-#28, #3-#27, #3-#29, #3-23-#5, #33-pSB1AK3, #34--pSB1AK3, #35--pSB1AK3)
- Transformation(ligation products:WT, #2-9:cheZ-, #14, #17) -> 37 incubation
'11/08/16(Tue)
- Colony PCR(#3-#14-5, #24-#3-17-5, #20-#28, #3-#27, #3-#29, #3-23-#5)
- Culture (#20-#28, #5, #14, #17, #2-9, #2-4, #2-3-17-5, #1, #2)
- Incubation (#24-#3-11-5, #33-pSB1AK3, #34-pSB1AK3, #35-pSB1AK3)
- Colony PCR(additional)(previously incubated ligation products: 3, 8, 10)
- hotstart PCR(sulAp, uvrAp, uvrBp)
- Digest(SPcut: #3, Ecut:#3(control)) go to 37 incubation
- Ligation retry(#3-#14-5, #3-#27, #3-#29, Negative control :#3)
- Frozen stock(#14, #17)
- Culture test(#14 on 0.3%Agar.(km))
- Ethanol precipitation: 29.9ng/ul * 1000ul
- Miniprep(Ligation products : #2-3-17-5, #14, #17, #1, #2)
- Electrophoresis(Colony PCR products: 3, 8, 10)
- Waking from frozen stock(#3, CheZ-, WT)
'11/08/17(Wed)
- Hotstart-PCR (Change conditions a little)
- From Genome
- From nested-region
- From parts
- Making Competent Cell, SOB medium
- Ethanol precipitation(WT: 40ng/ul)
- Culture from master-plate(#6-5, #20-28, #3-27, #33, #24-3-11-5 #24-3-17-5) -> Miniprep
- Assay(IPTG induced cell lysis device
- Gel extraction(SPcut:#3)
- Culture from Frozen stock(#1, #2, #2-3-17-5)
- Electrophresis check(Loading only DNA size markers)
- Miniprep(#3, #6-5, #20-28, #3-27, #33, #24-3-11-5 #24-3-17-5)
'11/08/18(Thu)
- Digest(8/17#3, #33 SPcut, #14-5 XPcut, #33 Ecut, #3-27,#3-30,#20-28 EScut, pSB1K3 EPcut)
- Hotstart-PCR (Change conditions a little)
- From Genome : uvrBp, cheZ
- From nested-region : uvrA
- Culture from 0809 master-plate ⑪ (#2-3-11-5) → Miniprep.
- Making LB plate & tube with various agarose conc. (plate: 0.5, 0.7, 0.9%×2 each, tube: 0.5,1.0,1.5,2.0,3.0%×2 each)(selection: Kanamycin)
- Gel extraction(8/17#3_SP,#33_SP, #14-5_XP, #33_E, #3-27_ES, pSB1K3 _EP)
- Miniprep(#1, #2, #2-3-11-5, #2-3-17-5)
- Digest(#1,#2 SPcut, #2-3-17-5 XPcut, #2-3-11-5 SPcut(check), EPcut(selection), #3-30, #20-28 EScut, 0817 Gel extracted PCR product (uvrBp) EScut, 0818PCR products (uvrAp, uvrBp, cheZ) ES&XPcut)
- Ligation (8/17#3-#14-5, 8/17#3-8/15#29, #33-#3-11-5, 8/15#34-pSB1AK3, #3-27-#5, #20-28-pSB1AK3, #28-pSB1AK3)
- Culture for frozen stock in eppendolf tube
'11/08/19(Fri)
- ColonyPCR(from ligation products)(#3-14-5, #3-29, #33-3-11-5, #34-pSB1AK3, #3-27-5)
- Gel extraction(08/18's digest products:#1_SP, #2_SP, #2-3-11-5_SP, #2-3-17-5_XP, cheZ_XP, #2-3-11-5_EP, 8/18uvrBp_ES, 8/17uvrBp_ES, uvrAp_ES)
- Colony PCR retry(#3-29, #34-pSB1AK3)
- Frozen Stock
- Check the list of frozen stock
- Assay of recAp from biobrick parts using RFP construct & transilluminator
- Electrophoresis(#3, #3-27, #3-30, #6-5, #7, #20-28, #24-3-11-5, #24-3-17-5, #33)
'11/08/22(Mon)
- Making SOB medium
- Culture for Miniprep
- From frozen stock(#1, #2, #7, #2-3-17-5(2), #3-30)
- From master plate(#3-14-5, #33-3-11-5, #34-pSB1AK3, #3-27-5)
- From master plate retry( at 13:50 OD600=0)
- From ligation products (for mistake above)
- Miniprep, Nanodrop and Electrophoresis
- From frozen stock(#1, #2, #2-3-17-5(2), #3-30) #7 was a little bit weak
- From ligation products(#7, #3-14-5, #3-27-5, #34, #33-3-11-5)
- Digest
- From Frozen Stock (#7_EX, #30_XP, #2-3-17-5_XP, #3-30_ES)
- From Master plate(#3-14-5_XP, #34-pSB1AK3_SP, #3-27-5_XP)
- Sulvage #33 using protocol of transformation
- Culture for Frozen stock
- From Master plate(#24-3-17-5, #14-5), 700ul, in eppendolf tube
'11/08/23(Tue)
- Gel extraction
- from O.N. digest products(#7_EX, #30_XP, #2-3-17-5_XP, #3-30_ES, #3-14-5_XP, #34-pSB1AK3_SP, #3-27-5_XP)
- Frozen stock
- From O.N. culture in eppendolf tube 700ul(#24-3-17-5, #14-5)
- From O.N. culture(for Miniprep)(#34, #3-27-5, #33-3-11-5)
- Miniprep
- retry from master plate #3-14-5(278.1 ng/ul), #33-3-11-5(188.9ng/ul), #34-pSB1AK3(89.6ng/ul), #3-27-5(327.0 ng/ul) O.N. culture
- Digest (#14-5 XPcut, #3-27-5 Xcut, Pcut (check), #3-27-5 EScut (sulvage) )
- Ligation (#14-#5, #3-30-#5, #3-#30, #34-#3-11-5 #20-#28)
- #33 sulvage (isolation culture)
- Making culture(LB amp, LB)
'11/08/24(Wed)
- Gel extraction & Electrophoresis(#3-27-5_ES, #14-5_XP, #3-27-5_X, *PCR for amplification (or cloning) -> PCR clean-up system -> digest
- #27 (82.1 ng/ul, total = 30 ul) -> XP cut
- #28 (104.4 ng/ul, total = 30 ul) -> XP cut
- Making plate and amp stock ->4℃
- amp 32
- km 11 (km conc. = 3/4 * that it was)
- Miniprep and Nanodrop
- #33 32.1 ng/ul <- there is probability that culture volume was not enough ->transfromation
- Luciferase assay reagents prep
- D-Luciferin Solution : 1ml*10 + 100ul*10 at -80℃
- Coelenterazine Solution : 100ul * 10 at -20℃
- Lysis buffer : 500ul * 10 at -20℃
- Ligation(#3-30-#5, #3-11-5-#34, #30-#3, #28-#20, #35-pSB1AK3, #2-3-11-5-pSB1K3, #14-5-#3 and Negative control)
'11/08/25(Thu)
- Renilla luciferase assay (with #2-3-17-5 culture)
- Gel extraction (#27, #28 XPcut)
- Ligation (0824 retry & #20-#28, #3-#27)
- Transformation (Ligation products into WT competent cells & #14-5 miniprep product into WT cell (line-drawing spread), #33 miniprep product into ΔcheZ cell (line-drawing spread))
- Digest (#14 XPcut)
- Passage(#2-3-17-5, WT, ΔcheZ, WT in eppen, ΔcheZ in eppen)
'11/08/26(Fri)
- Renilla luciferase assay (with #2-3-17-5 culture)
- Colony PCR (#33 Miniprep transformations)
- Gel extraction (#14_XP)
- Ligation (retry) (using today's competent cells, 0802 ΔcheZ cells)
- Cloning (cheZ)
- Digest (#5 EXcut, cheZ PCR products XPcut)
- Passage in a shaker(#2-3-17-5, WT, ΔcheZ)
- Making competent cell, LB broth
- Contamination check (0817 comp., 0802 comp., 0802 ⊿cheZ comp. spread on LB (amp) plates)
'11/08/27(Sat)
- Ligation & Transformation (0826 ①②③⑤⑧⑨) (into Nippon Gene comp.)
- Contamination check (WT FS, ΔcheZ Frozen stock)
- Waking from Frozen stock(WT, ΔcheZ)
'11/08/28(Sun)
- Making ΔcheZ comp., LB amp plates, TB
- Colony PCR & master plate preparation -> Failure!!
- Gel extraction (#29_XP, #5_EX)
- Miniprep and frozen stocks (#33, #14-5)
- Digest (#20 SPcut, #14-5 XPcut)
- Waking from frozen stock (WT, ⊿cheZ, #20, #14-5)
'11/08/29(Mon)
- PCR purification -> digest with XP(#27, #28)
- Miniprep(#20, #14-5)
- Gel extraction(#14-5, #20)
- Ligation (higher I/V ratio, into cheZ comp.)(#3-#14-5(retry), #2-3-11-5 to pSB1K3 (retry), #35-pSB1AK3, (#33,#34)-#3-17-5)
'11/08/30(Tue)
- Making LB amp, Aspartic acid solution (10mM), H2O2 solution (10mM)
- Gel extraction -> PCR was failed!(#27,#28)
- There was trash (from PCR) below
- Colony PCR(#3-#14-5, #2-3-11-5 to pSB1K3, #35-pSB1AK3, (#33,#34)-#3-17-5)
- PCR -> check (5uL EP -> PCR clean-up(#27,#28)
- Digest(#20_SP, pSB1AK3_EP, #3_SP)
- EP for MP check(#14-5, #20)
- Ligation(#3-#29, pSB1AK3-#2-3-11-5 on Amp/Km plate, #3 (Negative control), pSB1AK3 (Negative control) on Amp/Km plate
- Transformation(#2-9 into cheZ-) -> Line-drawing plating
- SOS stress (UV irradiation or H2O2)(#24-3-11-5, #33-3-11-5)
- culture for Miniprep from master plate(#34-3-17-5)
- Waking from frozen stock(#24-3-11-5, #33-3-11-5)
'11/08/31(Wed)
- Making LB plates(amp, km)
- Luciferin reagent preparation
- Flask A.C. for making competent cells
- Miniprep (#34-3-17-5)
- Gel extraction(#20_SP, pSB1AK3_EP, #3_SP)
- PCR -> clean-up -> XPcut(#27, #28, #14-5, #3-30)
- Waking from frozen stock(#2-9 (WT), #24-3-17-5)
- Plating from liquid culture(#2-3-17-5)
- Ligation(#20-#3-17-5, #2-3-11-5-pSB1AK3, #3-#14, #20-#28, #3-#27, #3-#29, #35-pSB1AK3, #3-#14-5, #3-30-#5, #33-#3-17-5 & Negative control)
- Transformation
- Plasmid isolation(#34-3-17-5(MP;1:9gradient))
- Part cloning(#36, #37)
- Contamination check(pSB1AK3(EPcut), pSB1K3(EPcut))
- Titer check(#3-17-5(Takara comp.,1:9gradient), #2-3-11-5(ΔcheZ comp.0802, 1:9gradient), #2-3-11-5(ΔcheZ comp.0830, 1:9gradient))
- Ligation products
September
'11/09/01(Thu)
- Colony PCR
- #34-3-17-5(MP), #3-14, #3-29, #35-pSB1AK3, #3-14-5, #37(from Part cloning)
- Gel extraction
- #27_XP ,#28_XP(0830); #27_XP, #28_XP, #3-30_XP -> failed!, #14-5_XP(0831)
- Making medium(SOB, LB amp plate), competent cells (WT & ΔcheZ)
- Miniprep(#3, #20)
- PCR and Column purification
- #3-14, #3-29, #3-14-5 (Recovery from colony PCR tubes!)
- Digest
- #3 -> Failed (Mixed with DMSO!), #20
- #3-14, #3-29, #3-14-5
- Ligation
- #3-27, #20-#3-17-5
- Transformation
- #36 into Nippon Gene comp.
- Ligation products into Takara comp. (including N.C.)
- Titer check(MP product #3 into 0901 WT comp., MP product #20 into 0901 ΔcheZ comp.)
- Mobility assay(#14-5 (km resistance) WT/ΔcheZ cells)
'11/09/02(Fri)
- Making medium and reagent(LB 120ml(0.125, 0.25, 0.5% Agar, M9 stock(20×), M9 200ml, Vitamins Solution (100×))
- Gel extraction -> Digest products were OK but extraction was failed.
- #20_SP, #3-14_XP, #3-14-5_XP, #3-29_XP
- Colony PCR & Master plate(#20-3-17-5, #3-27)
- Miniprep(#3, #20, #37, #3-14, #3-29, #3-14-5, #20-3-17-5, #3-27, ΔcheZ)
- Hot-start PCR & Column purification -> PCR products were OK but all purified products were lost.
- #20-3-17-5, #3-27 (Recovery from colony PCR tubes)
- #3-14, #3-14-5, #3-29 (from miniprep products)
- PCR & Column purification
- #20-3-17-5, #3-27, #3-14-5, #3-29 (from miniprep products)
- Digest
- #3_SP, #20_SP, #37_ES, #3-14_ES, #3-29_ES, #3-14-5_XP, ΔcheZ_E/P (from MP products)
- #20-3-17-5_XP, #3-27_ES, #3-14-5_XP, #3-29_ES (from PCR products)
'11/09/03(Sat)
- Gel extraction
- Ligation & Transformation
- #20-#3-14-5, #2-#3-14-5, #3-27-#5 into Nippon Gene comp. (+ High Competent Broth 240ul)
- #3-27-#5, #37-#5 into 0902 WT comp.(+ Takara SOC broth 500ul)
- Assays
- Mobility assay (.5%, .25%, .125%)
- Asp assay
- Part Cloning (#36)
'11/09/04(Sun)
- Hot-start PCR Pre-Mix (4×) preparation (Buffer + dNTPs + THUNDER Taq)
- Colony PCR(#20-3-14-5, #3-27-5, #3-29-5, #37-5)
- Colony Diffusion Assays
- GFP Live-Imaging Assays
- Ligation & Transformation (retry)
- #2-#3-14-5, #3-27-#5, #3-29-#5, #37-#5, #20-#28 into 0901 WT comp.
- #36 into Takara comp.
- #2, #5 as Negative Controls into 0901 WT comp.
- Culture for Miniprep(#20-3-14-5, #20)
- Digest(#20)
'11/09/05(Mon)
- Colony Diffusion Assays (#2-9) WT,cheZ-
- Gel extraction(#20)
- Luciferase Assays
- Calibrations (Firefly Luciferase standards)
- Background Assay
- Dual Luciferase Assay(#20-3-14-5, #20-3-17-5)
- Colony PCR(#2-#3-14-5, #3-27-#5, #3-29-#5, #37-#5, #20-#28, #36)
- PCR -> clean-up -> Digest(#3-29-#5, #37-5)
- IPTG-inducted Cell lysis test(IPTG+, LB plate (Positive control))
- Miniprep(#20-3-14-5, #37-5, #36)
- Making M9 -> Asp -> all failed(.125%, .25% Agar)
- Culture for Miniprep(#2-3-14-5)
'11/09/06(Tue)
- Gel extraction(#37-5, #3-29-5)
- Miniprep(#2-3-14-5, #20-3-14-5, #20-3-17-5, #36, #3-29-5, #37-5)
- PCR (retry) (Hot-start & Normal PCR) -> Purification -> Digest(#3-29-5_XP, #37-5_ES)
- PCR (cloning retry)(lexA, uvrBp)
- Ligation & Transformation
- #3-27-#5, #3-30-#5, #2-3-11-5-pSB1AK3
- #5, pSB1AK3 (N.C.)
- Culture for GFP live-imaging (#2-9 WT/cheZ-) -> 4℃ storage
- Ninhydrin sol. preparation (Not dissolvable!) -> 4℃ storage
'11/09/07(Wed)
- Colony PCR(#3-27-5, #3-30-5)
- Digest(#36_SP)
- Culture from master plate(#36, #3-29-5, #37-5)
- Miniprep(#36, #3-29-5, #37-5, #3-27-5 (failed))
- PCR purification & Digest(lexA_XP, uvrBp_ES)
- Gel extraction(#3-29-5_XP, #37-5_ES, pSB1AK3_EP, #36_SP)
- PCR amplification -> Digest(#3-14-5, #3-29-5, #37-5, #3-27-5)
- Digest(#36_SP (re), #2_SP, #2-3-17-5_EX, #3-14-5_ES, #34_SP)
- Culture for Miniprep(#2, #34, #36, #3-27-5, #3-14-5, #2-3-17-5)
Ligation & Transformation
- #33-#3-14-5, #34-#3-14-5, #36-#3-14-5, #2-#3-29-5, #20-#3-29-5, #3-#37-5, #2-3-11-5-pSB1AK3, #1-#3-17-5
- #33, #34, #36, #1, #2, #20, #3, pSB1AK3 (N.C.)
- Ninhydrin stock solution preparation
- Asp conc. check (it took 5 min at 80C)
- Making M9 medium(.25, .125% Agar.)
'11/09/08(Thu)
- Making competent cell (WT), LB(amp) plate
- Frozen stocks(#36, #3-14-5, #3-27-5)
- Colony PCR(#33-3-14-5, #34-3-14-5, #36-3-14-5, #20-3-29-5, #2-3-11-5(psB1AK3))
- Miniprep(#2, #34, #36, #2-3-17-5, #3-27-5, #3-14-5)
- Gel extraction
- PCR product: #35_SP, #31_XP, #3-14-5_XP, #3-27-5_XP, #3-29-5_XP, #37-5_XP
- Miniprep product: #2_SP, #34_SP, #36_SP, #2-3-17-5_EX, #3-14-5_ES
- PCR amplification(#34-3-14-5, #36-3-14-5, #20-3-29-5, #20-3-14-5)
- PCR for Sequencing -> Store at -20C(#33-3-11-5, #34-3-14-5, #20-3-29-5, #37-5)
- Ligation(#35_ES-psB1AK3_ES, #31_XP-psB1A2_XP, #31_XP-psB1AK3_XP, #3-#37-5, #36-3-29-5, #3-14-5_ES-#2-3-17-5_EX, #20-#3-27-5)
- Culture for Miniprep(#34-3-17-5, #36-3-14-5, #20-3-29-5, #2-3-11-5)
- Digest
- PCR products: #34-3-14-5_ES, #36-3-14-5_ES, #20-3-14-5_ES
- pSB1C3_EP
- Asp Gradient test
'11/09/09(Fri)
- Making competent cell (WT), TB
- Colony PCR(#35-pSB1AK3 (failed), #31-pSB1AK3, #3-#37-5, #36-#3-29-5, #3-14-5-#2-3-17-5, #20-3-27-5(failed))
- PCR amplification
- From Miniprep : #3-14-5, #3-17-5
- From Colony PCR : #31, #3-37-5, #36-3-29-5, #3-14-5-2-3-17-5 (failed)
- Digest
- Miniprep product: #3-11-5_EP
- PCR-purified product: #3-14-5_EP, #3-17-5_EP, #31_EP, #3-37-5_XP, #36-3-29-5_ES
- Miniprep(#34-3-14-5, #36-3-14-5, #20-3-29-5, #2-3-11-5 (pSB1AK3))
- Gel extraction(#34-3-14-5_ES, #36-3-14-5_ES, #20-3-14-5_ES, #3-11-5_EP)
- Ligation
- #3-11-5_EP-pSB1C3_EP, #29_XP-pSB1AK3_XP
- #36-3-14-5_ES-#2-3-17-5_EX, #20_SP-#3-27-5_XP(retry)
- Comp. check
- #2-3-11-5 (pSB1AK3) into 0908 WT comp.
- #20-3-29-5 into 0909 WT comp.
- #20-3-29-5 into 0802 cheZ- comp.
- Part cloning(BBa_J04450 (IPTG-induced RFP reporter on pSB1C3))
- Culture for Miniprep(#31-pSB1AK3, #3-37-5, #36-3-29-5, #3-14-5-2-3-17-5)
- UV irradiation -> Luciferase Assay(#24-3-17-5, #34-3-17-5)
- Asp Gradient test
- CleanSEQ -> Sequencing(#33-3-11-5, #34-3-14-5, #20-3-29-5, #37-5)
'11/09/10(Sat)
- Culture plate storage (4℃)
- Miniprep(#31-pSB1AK3, #3-37-5, #36-3-29-5, #3-14-5-2-3-17-5)
- Gel extraction(#3-14-5_EP, #3-17-5_EP, #31_EP, #3-37-5_XP, #36-3-29-5_ES)
- Colony PCR(#36-3-14-5-#2-3-17-5 (Maybe Ligation was FAILURE , estimated length is 1000-1500 by electrophoresis))
- Ligation(#20_SP-#3-37-5_XP)
'11/09/11(Sun)
- Digest from Miniprep products
- #3-14-5-2-3-17-5_XP
- #36-3-14-5_SP, #20_SP -> O.N.
- Colony PCR
- #3-11-5 (pSB1C3), #29 (pSB1AK3), BBa_J04450 (pSB1C3)
- #20-#3-27-5(failed), #20-#3-37-5, #36-#3-14-5-2-3-17-5
- Gel extraction(#3-14-5-2-3-17-5_XP)
- Culture for Miniprep
- From Master plate : #20-3-37-5, #29 (pSB1AK3), BBa_J04450 (pSB1C3)
- From plate stock : #20
- From frozen stock : #2-3-17-5
- Miniprep( #36-3-14-5-2-3-17-5)
- Ligation(#20_SP-#3-14-5-2-3-17-5_XP, #1_SP-#3-14-5-2-3-17-5_XP)
- Direct Ligation
- i.PCR amplification -> Digest for >1h -> Heat kill for 20min (#20-3-37-5_XP, #36-3-14-5-2-3-17-5_ES)
- ii.Digest for >1h -> Heat kill for 20min (#36-3-14-5-2-3-17-5_SP, pSB1A3_EP, #2-3-17-5_EX)
- iii.Ligation for 30min (#20-3-14-5_ES-#2-3-17-5_EX, #36-3-14-5-2-3-17-5_SP-#20-3-37-5_XP, #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1A3_EP)
- iv.Transformation
- Making 0.125% agar LB amp plate (IPTG 1, 10, 30, 100uM)
- Cell Diffusion Assay
- #20-3-29-5 WT (IPTG 1, 10, 30, 100uM)
- #20-3-29-5 cheZ-/- (IPTG 1, 10, 30, 100uM)
- WT
- cheZ-/-
'11/9/12 (Mon)
- Colony PCR
- #20-#3-14-5-2-3-17-5
- #20-3-14-5-#2-3-17-5
- #1-#3-14-5-2-3-17-5
- #36-3-14-5-2-3-17-5-#20-3-37-5 (no colony)
- #36-3-14-5-2-3-17-5-#20-3-37-5-pSB1A3 (PCR failed?)
Note: #20-3-37-5 colonies on the master-plate are RED. Maybe #2-3-11-5...
Note: #36-3-14-5-2-3-17-5-#20-3-37-5-pSB1A3 PCR amplification doesn't work. Seems failed extension...
- PCR amplification
- From master plate : #20-3-37-5
- From 09/11 Miniprep product : #36-3-14-5-2-3-17-5
Note: Only one #20-3-37-5 colony is NOT RED.
- Miniprep(#20, #29 (pSB1AK3), #2-3-17-5, #2-3-37-5, BBa_J04450 (pSB1C3))
- Digest
- From PCR : #36-3-14-5-2-3-17-5_ES, #20-3-37-5_XP
- pSB1A3_EP, pSB1C3_EP
- From Miniprep : #20-3-37-5_XP
- J04450_EP, *#2-9_XP, #20-3-29-5_ES
- Gel extraction
- #20_SP, #36-3-14-5_SP
- From OverNight digests : #2-3-17-5_EX
- #36-3-14-5-2-3-17-5_ES
- From today's PCR digests : #20-3-37-5_XP
- Culture for Miniprep(#20-3-37-5 from 0911 master plate)
- Dual Luciferase Assay(#20-3-14-5-2-3-17-5, #1-3-14-5-2-3-17-5, #36-3-14-5-2-3-17-5-20-3-37-5-pSB1A3)
- Ligation
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1A3_EP
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3_EP
- #3-27-5_XP-#20_SP
- #36-3-29-5_ES-#20-3-37-5_XP-pSB1AK3_EP
'11/9/13 (Tue)
- Colony PCR
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1A3_EP
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3_EP (failed)
- #3-27-5_XP-#20_SP (failed)
- #36-3-29-5_ES-#20-3-37-5_XP-pSB1AK3_EP
- Culture for Luciferase Assay
- #36-3-14-5-2-3-17-5-20-3-37-5(pSB1A3, pSB1C3) (IPTG 0, 1, 10, 38, 100uM)
- #20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM)
- #1-3-14-5-2-3-17-5 (IPTG 0uM)
Note: Seemes OD600 should NOT exceed 0.4~0.5, or intracellular luciferase will be saturated.
- Miniprep.
- #20-3-14-5-2-3-17-5
- #1-3-14-5-2-3-17-5
- #20-3-37-5 (from a non-red colony on the master plate)
- Gel extraction
- #20-3-37-5_XP (from non-red colony)
- #20-3-29-5_ES(failed)
- #2-9_XP, pSB1C3_EP
Note: #20-3-29-5 was not cut!
- Dual Luciferase Assay
- #1-3-14-5-2-3-17-5 (IPTG 0uM)
- #20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM)
- #36-3-14-5-2-3-17-5-20-3-37-5 (#2-3-11-5?) (IPTG 0, 1, 10, 38, 100uM)
- Sequencing preparation
- #20-3-37-5 (from a non-red colony on the master plate)
- #20-3-29-5
- #3-27-5
- #31
- Digest
- #20_E
- #3-29-5_S (from MP products) (cut check)
- Ligation
- #20_SP-#3-27-5_XP
- Transformation
- #3-14-5-2-3-17-5
- #1-3-14-5-2-3-17-5
- #20-3-14-5-2-3-17-5 (for plate stock)
- #20-3-37-5 (from a non-red colony on the master plate) (for plasmid check)
'11/09/14(Wed)
- Colony PCR
- #3-27-5_XP-#20_SP (failed)
- Culture for Luciferase Assay (from plate stocks)
- #20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM)
- #1-3-14-5-2-3-17-5 (IPTG 0uM)
- Digest
- #3-14-5, #3-17-5, #3-14-5-2-3-17-5, #1-3-14-5-2-3-17-5, #20-3-14-5-2-3-17-5 (EPcut)
- #20_SP
- Culture for Miniprep.
- From plate stock : #20, #20-3-14-5-2-3-17-5, #3-14-5-2-3-17-5, #3-17-5
- From 09/11 master plate : #3-11-5 (pSB1C3)
- Part cloning(#19: BBa_R0010)
- Ligation
- #20_SP-#3-29-5_XP
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3
- Making WT competent cell
- Dual Luciferase Assay
- #1-3-14-5-2-3-17-5 (IPTG 0uM)
- #20-3-14-5-2-3-17-5 (IPTG 0, 1, 10, 38, 100uM)
- Asp TLC Assay
- Asp Diffusion Assay (0.25% Agar)
- Fumaric Acid (Fum) Sol. preparation
- Sequencing
- ①F⑤R#20-3-37-5 (from a non-red colony on the master plate)
- ②F⑥R#20-3-29-5 (#2-3-11-5?)
- ③F⑦R#3-27-5
- ④F⑧R#31
'11/09/15(Thu)
- Colony PCR(#19, #36-3-14-5-2-3-17-5-20-3-37-5 (failed), #20-3-29-5)
- Miniprep(#20, #20-3-14-5-2-3-17-5, #3-14-5-2-3-17-5, #3-17-5)
- Gel extraction
- #3-14-5(failed), #3-17-5, #3-14-5-2-3-17-5, #1-3-14-5-2-3-17-5, #20-3-14-5-2-3-17-5 (EPcut); #20_SP
- Making WT competent cell
- Making LB plates(amp:39, cm:18)
- Culture for Miniprep(#3-11-5 (pSB1C3), #20-3-29-5, #19)
- Ligation
- sulAp(ES)-#3-14-5-2-3-17-5_XP-pSB1C3_PCR digest(EP)
- recNp(ES)-#3-14-5-2-3-17-5_XP-pSB1C3_PCR digest(EP)
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3_PCR digest(EP)
- #3-27-5_XP-#20_SP
- #3-17-5_EP-pSB1C3_PCR digest(EP)
- #3-14-5_EP-pSB1C3_PCR digest(EP)
- #3-14-5-2-3-17-5_EP-pSB1C3_PCR digest(EP)
- #20-3-14-5-2-3-17-5_EP-pSB1C3_PCR digest(EP)
- Culture for Dual Luciferase Assay(#20-3-14-5-2-3-17-5)
- TLC check (LB, M9)
- Cell Diffusion Assay
- Racing Assay (0.25% Agar, 10mM Asp Sol.)
- Fumaric Acid (Fum) Sol. preparation (failed!!)
- Sequencing
'11/09/16(Fri)
- Colony PCR
- sulAp(ES)-#3-14-5-2-3-17-5_XP-pSB1C3_PCR digest(EP),
- recNp(ES)-#3-14-5-2-3-17-5_XP-pSB1C3_PCR digest(EP),
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3_PCR digest(EP),
- #3-27-5_XP-#20_SP,
- #3-17-5_EP-pSB1C3_PCR digest(EP),
- #3-14-5_EP-pSB1C3_PCR digest(EP),
- #3-14-5-2-3-17-5_EP-pSB1C3_PCR digest(EP),
- #20-3-14-5-2-3-17-5_EP-pSB1C3_PCR digest(EP)
- #36-3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3_PCR digest(EP) (retry; non-red colonies)
- Miniprep(#3-11-5 (pSB1C3), #20-3-29-5, #19)
- PCR amplification(#3-27-5, #36-3-14-5-2-3-17-5, pSB1C3)
- Sequencing prep(#20-3-29-5)
- Making agarose gel, WT competent cell,
- Electrophoresis check(pSB1C3_PCR)
- Digest(pSB1C3_ES, pSB1C3_EP, #3-27-5_XP, #36-3-14-5-2-3-17-5_ES)
- Dual luciferase assay
- #20-3-14-5-2-3-17-5 (1h, 1.5h culture)(IPTG 0, 1, 10, 100uM)(2.5mL)
- Fumaric Acid (Fum) Sol. preparation
- TLC test
- Asp diffusion test
- Asp racing assay
'11/09/17(Sat)
- Colony PCR (retry)
- #3-14-5, #3-17-5 (pSB1C3)
- #36-3-14-5-2-3-17-5-20-3-37-5
- Culture for Miniprep(#3-14-5, #3-17-5 (pSB1C3))
- Ligation(sulAp-pSB1C3, recNp-pSB1C3, #20-#3-27-5)
- Transformation(#20-3-29-5 into cheZ-)
- Making medium(.25% Agar. M9& LB each 20 plates)
- Cell diffusion assay
'11/09/18(Sun)
- Colony PCR & Culture for Miniprep(#36-3-14-5-2-3-17-5-20-3-37-5, #33-3-14-5-2-3-17-5)
- Miniprep(#3-14-5, #3-17-5 (pSB1C3))
- Ligation(#20-#3-27-5, recNp-#3-14-5-2-3-17-5, sulAp-pSB1C3, recNp-pSB1C3, #31-pSB1C3)
- Cell diffusion assay
- Innoculate colony(WT / cheZ-/- / cheZ-/- + cheZ plasmid)
- GFP imaging assay(Culture cells expressing GFP)
'11/09/19(Mon)
- Culture for Dual luciferase assay(#36-3-14-5-2-3-17-5-20-3-37-5, #33-3-14-5-2-3-17-5, #1-3-14-5-2-3-17-5, #20-3-14-5-2-3-17-5)
- Miniprep(#36-3-14-5-2-3-17-5-20-3-37-5, #33-3-14-5-2-3-17-5)
- Ligation(#24-#3-14-5-2-3-17-5, #20-#3-27-5)
- Sequencing(#36-3-14-5-2-3-17-5-20-3-37-5, #33-3-14-5-2-3-17-5, #20-3-29-5)
- Dual luciferase assay(#1~, #20~, #36~, #33~ )
- Asp racing assay(Innoculate colonies)
- GFP imaging assay
'11/09/20(Tue)
- colony PCR(20-3-27-5, #24-3-14-5-2-3-17-5)
- Gel extraction(#3-27-5_XP)
- PCR amplification -> Digest(#29_EP, #20-3-29-5_EP, #3-29-5_EP)
- Digest(#20_SP)
- Sequencing prep.
- Ethanol precipitation (10ul TE)
- #3-11-5(pSB1C3): 144.5ng/uL
- #3-14-5: 728.0ng/uL
- #3-17-5(pSB1C3): 505.7ng/uL
- Ligation(#31_EP-pSB1C3_EP, #36-3-29-5_ES-#20-3-37-5_XP-pSB1C3_EP)
- Luciferase assay
- Asp diffusion assay
'11/09/21(Wed)
- Making plates
- Gel extraction(#20_SP, #29_EP, #3-29-5_EP, #20-3-29-5_EP)
- Ligation
- #20_SP-#3-27-5_XP, #24_SP-#3-14-5-2-3-17-5_XP, #29_EP-pSB1C3_EP, #3-29-5_EP-pSB1C3_EP, #20-3-29-5_EP-pSB1C3_EP, #3-14-5-2-3-17-5_EP-pSB1C3_EP
- Transformation
- #36 from BioBrick? part
- #33-3-14-5-2-3-17-5 into BL21 comp.
- Digest(#3-14-5-2-3-17-5_SP, #3-14-5-2-3-17-5_ES, #3-29-5_ES)
- Luciferase Assay(#36-3-14-5-2-3-17-5)
- Cell diffusion assay
'11/09/22(Thu)
- Ligation (retry)
- #3-27-5_XP-#20_SP, #3-14-5-2-3-17-5-#24_SP, #29_EP-pSB1C3_EP, #3-29-5_EP-pSB1C3_EP, #20-3-29-5_EP-pSB1C3_EP, #3-14-5-2-3-17-5_EP-pSB1C3_EP
- Colony PCR(#36(->culture for mini prep), #33-3-14-5-2-3-17-5(->Maybe the amplification was failed))
- Gel extraction(#3-14-5-2-3-17-5_SP, _ES; #3-29-5_ES)
- Making plates
- M9 0.25% Agar. +IPTG +Amp.: 20
- LB Cm.: 20
- Ligation
- #3-14-5-2-3-17-5_SP-#20-3-37-5_XP,
- #3-14-5-2-3-17-5_ES-#20-3-37-5_XP-pSB1C3,
- #3-29-5_ES-#20-3-37-5_XP-pSB1C3_EP
'11/09/23(Fri)
- colony PCR
- Column purification
- Ligation
- PCR amplification
- Competent cell(BL21(DE3))
- Asp diffusion test
- Cell diffusion assay
- Asp chemotaxis assay
'11/09/26(Mon)
- Ligation(#20-#3-27-5, #24-#3-14-5-2-3-17-5, recNp-3-14-5-2-3-17-5)
- Digest(pSB1C3_ES)
- Miniprep(#3-29-5-pSB1C3, #20-3-29-5-pSB1C3, #36)
- PCR amplification(pSB1A3, #3-29-5)
- Ethanol precipitation(#3-29-5-pSB1C3, #20-3-29-5-pSB1C3)
'11/09/27(Tue)
- Colony PCR(#24-3-14-5-2-3-17-5, #20-3-27-5, #35-3-14-5-2-3-17-5)
- Digest(#3-29-5_ES, #36_SP, pSB1C3_ES, pSB1C3_XP)
- Ligation(#20-3-37-5_XP-#3-14-5-2-3-17-5_SP, pSB1C3_ES, #31_EP-pSB1C3_EP)
- Gel digestion(#3-29-5_ES, #36_SP, pSB1C3_ES, pSB1C3_XP)
- Nanodrop(#36_SP, pSB1C3)
- PCR amplification(J04450)
- Gel extraction(pSB1C3_ES)
'11/09/28(Wed)
- Nanofrop
- From PCR : #3-29-5, pSB1C3
- From Miniprep : #3-29-5, #20-3-27-5, J04450
- Transformation(#3-5-3-14-5-2-3-17-5) -> BL21(DE) comp.
- PCR amplification and Digest(EPcut) (#31, #20-3-27-5)
- Digest(pSB1C3_EP, pSB1C3_XP, #3-29-5_ES, #24_SP_SP, #24_SP)
- Sequencing(#20-3-27-5, #3-29-5, #3-14-5, #3-17-5)
- Making AspA culture
- Cell diffusion assay
- Asp taxis assay
- Gel extraction and Nanodrop(pSB1C3_EP, pSB1C3_XP, #3-29-5_ES, #24_SP)
- Ligation
- #3-29-5_ES-#20-3-37-5_XP-pSB1C3_EP
- #3-14-5-2-3-17-5_XP-pSB1C3_XP
- #20-3-37-5_XP-pSB1C3_XP
- #3-27-5_XP-pSB1C3_XP
- #29_XP-pSB1C3_XP
- #24_SP-#3-14-5-2-3-17-5_XP
- #36_SP-#3-29-5_XP
'11/09/29(Thu)
- Ligation(#20-3-27-5_EP-pSB1C3_EP)
- Gel extraction and Nanodrop(#31_EP, #20-3-27-5_EP)
- PCR amplification(pSB1C3, #3-14-5-2-3-17-5)
- Digest(pSB1C3_EP, #3-14-5-2-3-17-5_XP)
- Colony PCR, PCR amplification, Miniprep and Colony diffusion assay(#36_SP-#3-29-5_XP)
- Colony PCR and Miniprep(#24-3-14-5-2-3-17-5)
'11/09/30(Fri)
- Digest(#36-3-29-5_ES)
- PCR amplification and Digest(EPcut)(#3-14-5-2-3-17-5, #20-3-27-5, #29, #31)
- Digest(#3-14-5-2-3-17-5_EP, #36-3-29-5_ES, pSB1C3_EP, pSB1C3_ES)
- Culture for MIniprep from plate stock(#3-14-5-2-3-17-5)
- Making competent cell(JM109)
- Gel extraction and Nanodrop(#36-3-29-5_ES_PCR, #36-3-29-5_ES_Miniprep, #3-14-5-2-3-17-5_EP, pSB1C3_EP, pSB1C3_ES, #3-14-5-2-3-17-5_EP, #20-3-27-5_EP, #29_EP, #31_EP)
- Ligation(#3-14-5-2-3-17-5_EP-pSB1C3_EP, #20-3-27-5_EP-pSB1C3_EP, #29_EP-pSB1C3_EP, #31_EP-pSB1C3_EP, #36-3-29-5_ES-#20-3-37-5_XP-pSB1C3_EP, recNp_ES-pSB1C3_ES, sulAp_ES-pSB1C3_ES, #3-14-5-2-3-17-5_EP-pSB1C3_EP)
- Sequencing prepartion(#20-3-27-5_F, #36-3-29-5
- Miniprep, Digest(EXcut) and Nanodrop(#3-14-5-2-3-17-5)
- Culture for Miniprep(pSB1C3, #36-3-29-5)
- Making plate LB Cm
October
'11/10/01(Sat)
- Part Sequencing for parts shipping
- Miniprep(#36-3-29-5)
- Sequencing preparation and Colony PCR(#20-3-27-5_F, #36-3-29-5, #24-3-14-5-2-3-17-5)
- Colony PCR(#3-14-5-2-3-17-5)
- second colony PCR
'11/10/02(Sun)
- PCR amplification(using Quick Taq), Digest(Epcut), Gel extraction and Nanodrop(#3-14-5-2-3-17-5)
- Gel extraction(#3-14-5-2-3-17-5_EX)
- Dual Luciferase assay(#24)
- #36-3-29-5 Assay
- #36-3-29-5-20-3-37-5 Assay
- Ligation(recNp_ES-#3-14-5-2-3-17-5_EX, #3-14-5-2-3-17-5_EP-pSB1C3_EP)
- Making agarose gel
- Asp degradation assay
'11/10/03(Mon)
- Miniprep, Nanodrop and PCR sequencing(#20-3-27-5, #29, #31, #36-3-29-5-20-3-37-5, recNp, sulAp)
- Colony PCR(#3-14-5-2-3-17-5-pSB1C3)
- Preparation for shipping
'11/10/04(Tue)
- Preparation for shipping(#3-11-5, #3-14-5, #3-17-5, #3-14-5-2-3-17-5, #20-3-27-5, #29, #3-29-5, #20-3-29-5, #36-3-29-5-20-3-37-5, #31, #33, #35)