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Team:UT-Tokyo/LabNote
From 2011.igem.org
(Difference between revisions)
| Line 241: | Line 241: | ||
*Planting Negative control | *Planting Negative control | ||
*Making frozen stock | *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></html> | ||
| Line 257: | Line 257: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/06/17(Fri)=== | ==='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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/06/21(Tue)=== | ==='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 | *Nanodrop again | ||
<html></div></html> | <html></div></html> | ||
| Line 271: | Line 271: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/06/22(Wed)=== | ==='11/06/22(Wed)=== | ||
| - | + | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">9</span></html, J31000) | |
*Making competent cell | *Making competent cell | ||
<html></div></html> | <html></div></html> | ||
| Line 286: | Line 286: | ||
*Digest(product of Miniprep 06/09 EPcut) | *Digest(product of Miniprep 06/09 EPcut) | ||
*Electrophoresis | *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), | *Making 1×TAE, agarose gel, LB medium, LB plate(amp), | ||
<html></div></html> | <html></div></html> | ||
| Line 302: | Line 302: | ||
==='11/07/01(Fri)=== | ==='11/07/01(Fri)=== | ||
*Digest(EPcut) | *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></html> | ||
| Line 308: | Line 308: | ||
==='11/07/05(Tue)=== | ==='11/07/05(Tue)=== | ||
*Electrophoresis(product of digest 07/01) | *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) | *Making antibiotic stock(1000×)(Km,Cm) | ||
<html></div></html> | <html></div></html> | ||
| Line 315: | Line 315: | ||
==='11/07/06(Wed)=== | ==='11/07/06(Wed)=== | ||
*Gel extraction (pre)(BBa_I712052) | *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 | *Making agarose gel | ||
<html></div></html> | <html></div></html> | ||
| Line 321: | Line 321: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/07(Thu)=== | ==='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） | *Making LB plate（Cm,Km） | ||
<html></div></html> | <html></div></html> | ||
| Line 329: | Line 329: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/08(Fri)=== | ==='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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/11(Mon)=== | ==='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></html> | ||
| Line 345: | Line 345: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/13(Wed)=== | ==='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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/14(Thu)=== | ==='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 | *Making LB+amp | ||
<html></div></html> | <html></div></html> | ||
| Line 362: | Line 362: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/15(Fri)=== | ==='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 | *Dispensing Ligation Buffer | ||
| - | + | *Frozen stock(<nowiki>#</nowiki><html><span class="part_num">11</span></html) | |
<html></div></html> | <html></div></html> | ||
| Line 380: | Line 380: | ||
*Making Master plate | *Making Master plate | ||
*Testing product of Miniprep | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/21(Thu)=== | ==='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 | *Testing product | ||
*Digest | *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 | *Making TB | ||
<html></div></html> | <html></div></html> | ||
| Line 395: | Line 395: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/22(Fri)=== | ==='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 | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/25(Mon)=== | ==='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×) | *Defrost Primer(200×, 10×) | ||
*Dispensing PCR Mix | *Dispensing PCR Mix | ||
| Line 414: | Line 414: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/26(Tue)=== | ==='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) | *Making reagent for TB(KOH, CaCl2, KCl, MnCl2) | ||
<html></div></html> | <html></div></html> | ||
| Line 424: | Line 424: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/27(Wed)=== | ==='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 | *Electrophoresis | ||
<html></div></html> | <html></div></html> | ||
| Line 430: | Line 430: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/07/28(Thu)=== | ==='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 | *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 | *Making TB bugger, LB plate, 0.3% LB plate | ||
<html></div></html> | <html></div></html> | ||
| Line 442: | Line 442: | ||
==='11/07/29(Fri)=== | ==='11/07/29(Fri)=== | ||
*Cloning(lexA, cheZ) | *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) | *Digest(lexA, cheZ PCR product XP cut) | ||
<html></div></html> | <html></div></html> | ||
| Line 453: | Line 453: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/01(Mon)=== | ==='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) | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/02(Tue)=== | ==='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 | *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) | *Making competent cell(cheZ-, Wild Type) | ||
*Miniprep | *Miniprep | ||
| Line 474: | Line 474: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/04(Thu)=== | ==='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 | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/05(Fri)=== | ==='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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/08(Mon)=== | ==='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 | *Making Gel | ||
*Digest | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/09(Tue)=== | ==='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>-</htmlpsB1A2, <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>-</htmlpsB1A2, <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>-</htmlpsB1A2, <nowiki>#</nowiki><html><span class="part_num">31</span>-</htmlpsB1A2, <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 | *Making LB plate | ||
<html></div></html> | <html></div></html> | ||
| Line 509: | Line 509: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/10(Wed)=== | ==='11/08/10(Wed)=== | ||
| - | + | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">27</span>-</htmlpSB1A2, <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>-</htmlpsB1A2, <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>-</htmlpsB1A2, <nowiki>#</nowiki><html><span class="part_num">31</span>-</htmlpSB1A2, <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) | *Cloning(lexA, cheZ) | ||
*Digest | *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 | *Defrost Primer | ||
<html></div></html> | <html></div></html> | ||
| Line 521: | Line 521: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/11(Thu)=== | ==='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) | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/12(Fri)=== | ==='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) | *Cloning(nested-PCR)(uvrAp, sulAp) | ||
*Check Electrophoresis | *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 | *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 | **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) | *Cloning(hotstart-PCR: uvrBp , cheZ) | ||
*Digest | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/15(Mon)=== | ==='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 | *Digest | ||
**XPcut: cheZ, uvrBp(PCR 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) | |
*Assay of Cell lysis construction(pLuc-Lysis device) inducted by IPTG | *Assay of Cell lysis construction(pLuc-Lysis device) inducted by IPTG | ||
*Making Lysis buffer, Agarose gel | *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>-</htmlpSB1AK3, <nowiki>#</nowiki><html><span class="part_num">34</span>--</htmlpSB1AK3, <nowiki>#</nowiki><html><span class="part_num">35</span>--</htmlpSB1AK3) | |
| - | + | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/16(Tue)=== | ==='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>-</htmlpSB1AK3, <nowiki>#</nowiki><html><span class="part_num">34</span>-</htmlpSB1AK3, <nowiki>#</nowiki><html><span class="part_num">35</span>-</htmlpSB1AK3) | |
*Colony PCR(additional)(previously incubated ligation products: 3, 8, 10) | *Colony PCR(additional)(previously incubated ligation products: 3, 8, 10) | ||
*hotstart PCR(sulAp, uvrAp, uvrBp) | *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 | *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) | *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></html> | ||
| Line 584: | Line 584: | ||
*Making Competent Cell, SOB medium | *Making Competent Cell, SOB medium | ||
*Ethanol precipitation(WT: 40ng/ul) | *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 | *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) | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/18(Thu)=== | ==='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) | *Hotstart-PCR (Change conditions a little) | ||
**From Genome : uvrBp, cheZ | **From Genome : uvrBp, cheZ | ||
**From nested-region : uvrA | **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) | *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>-</htmlpSB1AK3, <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>-</htmlpSB1AK3, <nowiki>#</nowiki><html><span class="part_num">28</span>-</htmlpSB1AK3) | |
*Culture for frozen stock in eppendolf tube | *Culture for frozen stock in eppendolf tube | ||
<html></div></html> | <html></div></html> | ||
| Line 609: | Line 609: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/19(Fri)=== | ==='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>-</htmlpSB1AK3, <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>-</htmlpSB1AK3) | |
*Frozen Stock | *Frozen Stock | ||
*Check the list of frozen stock | *Check the list of frozen stock | ||
*Assay of recAp from biobrick parts using RFP construct & transilluminator | *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></html> | ||
| Line 622: | Line 622: | ||
*Making SOB medium | *Making SOB medium | ||
*Culture for Miniprep | *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>-</htmlpSB1AK3, <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 master plate retry( at 13:50 OD600=0) | ||
**From ligation products (for mistake above) | **From ligation products (for mistake above) | ||
*Miniprep, Nanodrop and Electrophoresis | *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 | *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>-</htmlpSB1AK3_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 | *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></html> | ||
| Line 640: | Line 640: | ||
==='11/08/23(Tue)=== | ==='11/08/23(Tue)=== | ||
*Gel extraction | *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>-</htmlpSB1AK3_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 | *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 | *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>-</htmlpSB1AK3(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) | *Making culture(LB amp, LB) | ||
<html></div></html> | <html></div></html> | ||
| Line 654: | Line 654: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/24(Wed)=== | ==='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℃ | *Making plate and amp stock ->4℃ | ||
**amp 32 | **amp 32 | ||
**km 11 (km conc. = 3/4 * that it was) | **km 11 (km conc. = 3/4 * that it was) | ||
*Miniprep and Nanodrop | *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 | *Luciferase assay reagents prep | ||
**D-Luciferin Solution : 1ml*10 + 100ul*10 at -80℃ | **D-Luciferin Solution : 1ml*10 + 100ul*10 at -80℃ | ||
**Coelenterazine Solution : 100ul * 10 at -20℃ | **Coelenterazine Solution : 100ul * 10 at -20℃ | ||
**Lysis buffer : 500ul * 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>-</htmlpSB1AK3, <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>-</htmlpSB1K3, <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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/25(Thu)=== | ==='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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/26(Fri)=== | ==='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) | *Ligation (retry) (using today's competent cells, 0802 ΔcheZ cells) | ||
*Cloning (cheZ) | *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 | *Making competent cell, LB broth | ||
*Contamination check (0817 comp., 0802 comp., 0802 ⊿cheZ comp. spread on LB (amp) plates) | *Contamination check (0817 comp., 0802 comp., 0802 ⊿cheZ comp. spread on LB (amp) plates) | ||
| Line 703: | Line 703: | ||
*Making ΔcheZ comp., LB amp plates, TB | *Making ΔcheZ comp., LB amp plates, TB | ||
*Colony PCR & master plate preparation -> Failure!! | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/08/29(Mon)=== | ==='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>-</htmlpSB1AK3, (<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></html> | ||
| Line 720: | Line 720: | ||
==='11/08/30(Tue)=== | ==='11/08/30(Tue)=== | ||
*Making LB amp, Aspartic acid solution (10mM), H2O2 solution (10mM) | *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 | *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>-</htmlpSB1AK3, (<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></html> | ||
| Line 738: | Line 738: | ||
*Luciferin reagent preparation | *Luciferin reagent preparation | ||
*Flask A.C. for making competent cells | *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>-</htmlpSB1AK3, <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>-</htmlpSB1AK3, <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 | *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)) | **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 | **Ligation products | ||
<html></div></html> | <html></div></html> | ||
| Line 758: | Line 758: | ||
==='11/09/01(Thu)=== | ==='11/09/01(Thu)=== | ||
*Colony PCR | *Colony PCR | ||
| - | + | **<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>-</htmlpSB1AK3, <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 | *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) | *Making medium(SOB, LB amp plate), competent cells (WT & ΔcheZ) | ||
| - | + | *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 | *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!) | |
*Digest | *Digest | ||
| - | + | **<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><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 | *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 | |
*Transformation | *Transformation | ||
| - | + | **<nowiki>#</nowiki><html><span class="part_num">36</span></html into Nippon Gene comp. | |
**Ligation products into Takara comp. (including N.C.) | **Ligation products into Takara comp. (including N.C.) | ||
| - | + | **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></html> | ||
| Line 781: | Line 781: | ||
*Making medium and reagent(LB 120ml(0.125, 0.25, 0.5% Agar, M9 stock(20×), M9 200ml, Vitamins Solution (100×)) | *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. | *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. | *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 | *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 | *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></html> | ||
| Line 798: | Line 798: | ||
*Gel extraction | *Gel extraction | ||
*Ligation & Transformation | *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 | *Assays | ||
**Mobility assay (.5%, .25%, .125%) | **Mobility assay (.5%, .25%, .125%) | ||
**Asp assay | **Asp assay | ||
| - | + | *Part Cloning (<nowiki>#</nowiki><html><span class="part_num">36</span></html) | |
<html></div></html> | <html></div></html> | ||
| Line 809: | Line 809: | ||
==='11/09/04(Sun)=== | ==='11/09/04(Sun)=== | ||
*Hot-start PCR Pre-Mix (4×) preparation (Buffer + dNTPs + THUNDER Taq) | *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 | *Colony Diffusion Assays | ||
*GFP Live-Imaging Assays | *GFP Live-Imaging Assays | ||
*Ligation & Transformation (retry) | *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> | <html></div></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/05(Mon)=== | ==='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 | *Luciferase Assays | ||
**Calibrations (Firefly Luciferase standards) | **Calibrations (Firefly Luciferase standards) | ||
**Background Assay | **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)) | *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) | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/06(Tue)=== | ==='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) | *PCR (cloning retry)(lexA, uvrBp) | ||
*Ligation & Transformation | *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>-</htmlpSB1AK3 | |
| - | + | **<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 | *Ninhydrin sol. preparation (Not dissolvable!) -> 4℃ storage | ||
<html></div></html> | <html></div></html> | ||
| Line 851: | Line 851: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/07(Wed)=== | ==='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) | *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 | 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>-</htmlpSB1AK3, <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 | *Ninhydrin stock solution preparation | ||
**Asp conc. check (it took 5 min at 80C) | **Asp conc. check (it took 5 min at 80C) | ||
| Line 871: | Line 871: | ||
==='11/09/08(Thu)=== | ==='11/09/08(Thu)=== | ||
*Making competent cell (WT), LB(amp) plate | *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 | *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 | *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 | **pSB1C3_EP | ||
*Asp Gradient test | *Asp Gradient test | ||
| Line 890: | Line 890: | ||
==='11/09/09(Fri)=== | ==='11/09/09(Fri)=== | ||
*Making competent cell (WT), TB | *Making competent cell (WT), TB | ||
| - | + | *Colony PCR(<nowiki>#</nowiki><html><span class="part_num">35</span>-</htmlpSB1AK3 (failed), <nowiki>#</nowiki><html><span class="part_num">31</span>-</htmlpSB1AK3, <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 | *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 | *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 | *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 | *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)) | *Part cloning(BBa_J04450 (IPTG-induced RFP reporter on pSB1C3)) | ||
| - | + | *Culture for Miniprep(<nowiki>#</nowiki><html><span class="part_num">31</span>-</htmlpSB1AK3, <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 | *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></html> | ||
| Line 916: | Line 916: | ||
==='11/09/10(Sat)=== | ==='11/09/10(Sat)=== | ||
*Culture plate storage (4℃) | *Culture plate storage (4℃) | ||
| - | + | *Miniprep(<nowiki>#</nowiki><html><span class="part_num">31</span>-</htmlpSB1AK3, <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></html> | ||
| Line 925: | Line 925: | ||
==='11/09/11(Sun)=== | ==='11/09/11(Sun)=== | ||
*Digest from Miniprep products | *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><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><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 | |
| - | + | *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) | |
*Culture for Miniprep | *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 | *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 | **iv.Transformation | ||
*Making 0.125% agar LB amp plate (IPTG 1, 10, 30, 100uM) | *Making 0.125% agar LB amp plate (IPTG 1, 10, 30, 100uM) | ||
*Cell Diffusion Assay | *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 | **WT | ||
**cheZ-/- | **cheZ-/- | ||
| Line 953: | Line 953: | ||
==='11/9/12 (Mon)=== | ==='11/9/12 (Mon)=== | ||
*Colony PCR | *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>-</htmlpSB1A3 (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>-</htmlpSB1A3 PCR amplification doesn't work. Seems failed extension... | |
*PCR amplification | *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 | *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 | **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 | *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>-</htmlpSB1A3) | |
*Ligation | *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></html> | ||
| Line 987: | Line 987: | ||
==='11/9/13 (Tue)=== | ==='11/9/13 (Tue)=== | ||
*Colony PCR | *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><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><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)<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><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">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 extraction | *Gel extraction | ||
| - | + | **<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><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><html><span class="part_num">2</span>-<span class="part_num">9</span></html_XP, pSB1C3_EP | |
| - | + | 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><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) | |
| - | + | **<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><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><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">3</span>-<span class="part_num">27</span>-<span class="part_num">5</span></html | |
| - | + | **<nowiki>#</nowiki><html><span class="part_num">31</span></html | |
*Digest | *Digest | ||
| - | + | **<nowiki>#</nowiki><html><span class="part_num">20</span></html_E | |
| - | + | **<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><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><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 (for plate stock) | |
| - | + | **<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></html> | ||
| Line 1,029: | Line 1,029: | ||
==='11/09/14(Wed)=== | ==='11/09/14(Wed)=== | ||
*Colony PCR | *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) | *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 | *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. | *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 | *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 | *Making WT competent cell | ||
*Dual Luciferase Assay | *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 TLC Assay | ||
*Asp Diffusion Assay (0.25% Agar) | *Asp Diffusion Assay (0.25% Agar) | ||
*Fumaric Acid (Fum) Sol. preparation | *Fumaric Acid (Fum) Sol. preparation | ||
*Sequencing | *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></html> | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/15(Thu)=== | ==='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 | *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 WT competent cell | ||
*Making LB plates(amp:39, cm:18) | *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 | *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) | *TLC check (LB, M9) | ||
*Cell Diffusion Assay | *Cell Diffusion Assay | ||
| Line 1,086: | Line 1,086: | ||
==='11/09/16(Fri)=== | ==='11/09/16(Fri)=== | ||
*Colony PCR | *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, | *Making agarose gel, WT competent cell, | ||
*Electrophoresis check(pSB1C3_PCR) | *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 | *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 | *Fumaric Acid (Fum) Sol. preparation | ||
**TLC test | **TLC test | ||
| Line 1,112: | Line 1,112: | ||
==='11/09/17(Sat)=== | ==='11/09/17(Sat)=== | ||
*Colony PCR (retry) | *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) | *Making medium(.25% Agar. M9& LB each 20 plates) | ||
*Cell diffusion assay | *Cell diffusion assay | ||
| Line 1,123: | Line 1,123: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/18(Sun)=== | ==='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>-</htmlpSB1C3) | |
*Cell diffusion assay | *Cell diffusion assay | ||
**Innoculate colony(WT / cheZ-/- / cheZ-/- + cheZ plasmid) | **Innoculate colony(WT / cheZ-/- / cheZ-/- + cheZ plasmid) | ||
| Line 1,133: | Line 1,133: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/19(Mon)=== | ==='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) | *Asp racing assay(Innoculate colonies) | ||
*GFP imaging assay<html></div></html> | *GFP imaging assay<html></div></html> | ||
| Line 1,143: | Line 1,143: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/20(Tue)=== | ==='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. | *Sequencing prep. | ||
*Ethanol precipitation (10ul TE) | *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 | *Luciferase assay | ||
*Asp diffusion assay | *Asp diffusion assay | ||
| Line 1,160: | Line 1,160: | ||
==='11/09/21(Wed)=== | ==='11/09/21(Wed)=== | ||
*Making plates | *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 | *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 | *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 | *Cell diffusion assay | ||
<html></div></html> | <html></div></html> | ||
| Line 1,174: | Line 1,174: | ||
==='11/09/22(Thu)=== | ==='11/09/22(Thu)=== | ||
*Ligation (retry) | *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 | *Making plates | ||
**M9 0.25% Agar. +IPTG +Amp.: 20 | **M9 0.25% Agar. +IPTG +Amp.: 20 | ||
**LB Cm.: 20 | **LB Cm.: 20 | ||
*Ligation | *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></html> | ||
| Line 1,200: | Line 1,200: | ||
<html><div class="labDay"></html> | <html><div class="labDay"></html> | ||
==='11/09/26(Mon)=== | ==='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) | *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>-</htmlpSB1C3, <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>-</htmlpSB1C3, <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>-</htmlpSB1C3, <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>-</htmlpSB1C3) | |
<html></div></html> | <html></div></html> | ||
Revision as of 17:16, 2 October 2011
Lab Note
iGEM UT-Tokyo
Parts List
Each part is represented by s number (e.g. #20 for lac promotor) in this notebook. We have also used this number in our hand-written lab notebook. By pointing the mouse on the number in the construct, you can find out the details of the part.
| Number | Reg.ID | Content | Plasmid | Length (bp) |
|---|---|---|---|---|
| #1 | J23119 | c.Promoter (Strong) | pSB1A2 | 35 |
| #2 | J23118 | c.Promoter (Medium) | BBa_J61002 | 35 |
| #3 | B0032 | RBS | pSB1A2 | 13 |
| #5 | B0014 | d.Ter | pSB1AK3 | 95 |
| #9 | E0240 | RBS-GFP-d.Ter | pSB1A2 | 876 |
| #10 | E0040 | GFP | pSB1A2 | 681 |
| #11 | E1010 | RFP | pSB2K3 | 723 |
| #14 | I712019 | fLuc | pSB1AK8 | 1653 |
| #17 | J52008 | rLuc | pSB1AK3 | 936 |
| #20 | R0011 | lacP | pSB1A2 | 55 |
| #21 | C0012 | LacI | pSB1A2 | 1128 |
| #22 | I712074 | pT7 | pSB1AK8 | 46 |
| #23 | K145001 | T7 RNA Pol. | pSB1A2 | 2655 |
| #24 | J22106 | recAp | pSB1A2 | 192 |
| #27 | C0083 | AspA | pSB2K3 | 1518 |
| #28 | K112808 | T4 phage lysis device (no promoter) | pSB1A2 | 1785 |
| #29 | - | CheZ | pSB1AK3 | 728 |
| #30 | K117000 | Lysis gene | pSB1A2 | 144 |
| #31 | - | LexA | pSB1AK3 | 750 |
| #33 | - | sulAp | pSB1AK3 | 67 |
| #34 | - | uvrAp | pSB1AK3 | 96 |
| #35 | - | recNp | ||
| #36 | R0051 | cI-repressed promoter | pSB1A2 | 49 |
| #37 | C0051 | cI repressor (LVA tagged) | pSB1A2 | \ |
| #38 | - | RecA |
Lab Diary
- 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#5#10
'11/06/14(Tue)
- Picking up colony
- Making Mg reagent
'11/06/15(Wed)
- Miniprep
'11/06/17(Fri)
- Picking up colony(#3#9#5#1#2#24
'11/06/21(Tue)
- Making frozen stock(#3#9#9
'11/06/22(Wed)
- Miniprep(#9
'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#17
'11/06/29(Wed)
- Picking up colony
- Making LB broth
July
'11/07/01(Fri)
- Digest(EPcut)
- Miniprep(BBa_E0030, E0020, #14#17
'11/07/05(Tue)
- Electrophoresis(product of digest 07/01)
- Digest(#9#14#17#3#5
'11/07/06(Wed)
- Gel extraction (pre)(BBa_I712052)
- Picking up colony(Ba_J23119, #2#24#10
'11/07/07(Thu)
- Digest（BBa_E0240, #14#17#3#5#20#21#22#23#20#21#22
'11/07/08(Fri)
- Miniprep(BBa_J23119, #2#10#24
'11/07/11(Mon)
- Gel extraction(#9#14#17#1#11#20#21#22#23
'11/07/12(Tue)
- Picking up colony
'11/07/13(Wed)
- Frozen stock(#1#20#21#14#23#11#3#5#2
'11/07/14(Thu)
- Miniprep(#1#11#20#21#22#23#2#2#3#5#11
'11/07/15(Fri)
- Ligation(#2-#9#2-#3#14-#5#17-#5#2-#9#2-#3#14-#5#17-#5#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#20#2-3#10#11#14-5#17-5
'11/07/22(Fri)
- Frozen stock(#2#2-3#2-9#14-5#17-5#20#2-3#10#11#14-5#17-5#20#10#11#14-5#17-5#27#28
'11/07/25(Mon)
- Colony check(#2#2-9#20#10#11#14-5#17-5#20-#9#3-#11#3-#14-5#3-#17-5
'11/07/26(Tue)
- Picking up colony and making master plate(#20-9#3-11#3-14-5#3-17-5#20-9#3-11#3-14-5#3-17-5#14-5#22#23#3-#14-5#3#15#27#28#30#3-14-5#3
'11/07/27(Wed)
- Miniprep(#1#2#3#10#22#24#20-9#3-11#3-17-5
'11/07/28(Thu)
- Gel extraction(#14-5#22#23#1#2#3#24#3-11#3-17-5#3-#14-5
'11/07/29(Fri)
- Cloning(lexA, cheZ)
- Colony PCR(#3-#14-5#1#2#3#24#3-11#3-17-5#30
August
'11/08/01(Mon)
- Nanodrop(#3-11#1#3#24#2#3-17-5#30#3#5#9#30#3#9#30#2-#3-17-5#3-11-#5#14-#5#3-#23#22-#9#3-#30#2-#3-17-5#3-11-#5#14-#5#3-#23#22-#9#3-#30#27#28
'11/08/02(Tue)
- Colony PCR(#2-#3-17-5#3-11-#5#14-#5#3-#23#22-#9#27#28#27#28#30
'11/08/04(Thu)
- Electrophoresis(product of ligation #2-3-17-5#3-11-5#14-5#22-9#2-3-17-5#3-11-5#14-5#22-9#23#20-9
'11/08/05(Fri)
- Digest(#14-5#3-11-5#27#28
'11/08/08(Mon)
- Defrost Frozen stock, Miniprep, Nanodrop and Electrophoresis(#5#20#23#30#27#28#3-11-5#23#30#6#7#7#20#5#14-5#3-11-5
'11/08/09(Tue)
- Miniprep, Electrophoresis and Digest(#5#14-5#3-11-5#6#27#8#3-11-5#23#7#20#7#27-#3-27#20-28#28-#3-29#29-#31-#3-#23#1-#3-11-5#2-#3-11-5#24-#3-11-5#3-#30
'11/08/10(Wed)
- Colony PCR(#27-#3-27#20-28#28-#3-29#29-#31-#3-#23#1-#3-11-5#2-#3-11-5#24-#3-11-5#3-#30#6#7#1#2#3#7#20#23
'11/08/11(Thu)
- Gel extraction(#30#14-5#5#3-11-5#3#20-28#31#24-3-11-5#3-30
'11/08/12(Fri)
- Colony PCR(#3-14-5#6-5#3-23#27#28#3-14-5#5#5#14#5#14-5#3-30#27#28#13-14-5#6-5#3-23#27#28#6-5#3-23
'11/08/15(Mon)
- Gel extraction(from previous digest products(8/12) : #27#28#33#34#4#3-23#3-11-5#3-11-5#3-#14-5#24-#3-17-5#24-#3-11-5#20-#28#3-#27#3-#29#3-23-#5#33-#34--#35--#2-9#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#20-#28#5#14#17#2-9#2-4#2-3-17-5#1#2#24-#3-11-5#33-#34-#35-#3#3#3-#14-5#3-#27#3-#29#3#14#17#14#2-3-17-5#14#17#1#2#3
'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#1#2#2-3-17-5#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#14-5#33#3-27#3-30#20-28#2-3-11-5#3#33#14-5#33#3-27#1#2#2-3-11-5#2-3-17-5#1#2#2-3-17-5#2-3-11-5#3-30#20-28#3-#14-5#3-8#29#33-#3-11-5#34-#3-27-#5#20-28-#28-
'11/08/19(Fri)
- ColonyPCR(from ligation products)(#3-14-5#3-29#33-3-11-5#34-#3-27-5#1#2#2-3-11-5#2-3-17-5#2-3-11-5#3-29#34-#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#3-30#3-14-5#33-3-11-5#34-#3-27-5#1#2#2-3-17-5#3-30#7#7#3-14-5#3-27-5#34#33-3-11-5#7#30#2-3-17-5#3-30#3-14-5#34-#3-27-5#33#24-3-17-5#14-5
'11/08/23(Tue)
- Gel extraction
- from O.N. digest products(#7#30#2-3-17-5#3-30#3-14-5#34-#3-27-5#24-3-17-5#14-5#34#3-27-5#33-3-11-5#3-14-5#33-3-11-5#34-#3-27-5#14-5#3-27-5#3-27-5#14-#5#3-30-#5#3-#30#34-#3-11-5#20-#28#33
'11/08/24(Wed)
- Gel extraction & Electrophoresis(#3-27-5#14-5#3-27-5 PCR clean-up system -> digest
**
# 27 XP cut **# 28 XP cut *Making plate and amp stock ->4℃ **amp 32 **km 11 (km conc. = 3/4 * that it was) *Miniprep and Nanodrop **# 33transfromation *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-#2-3-11-5-#14-5-#3
'11/08/25(Thu)
- Renilla luciferase assay (with #2-3-17-5#27#28#20-#28#3-#27#14-5#33#14#2-3-17-5
'11/08/26(Fri)
- Renilla luciferase assay (with #2-3-17-5#33#14#5#2-3-17-5
'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#5#33#14-5#20#14-5#20#14-5
'11/08/29(Mon)
- PCR purification -> digest with XP(#27#28#20#14-5#14-5#20#3-#14-5#2-3-11-5#35-#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#3-#14-5#2-3-11-5#35-#33#34#3-17-5 check (5uL EP -> PCR clean-up(
# 27#28#20#3#14-5#20#3-#29#2-3-11-5#3#2-9 Line-drawing plating *SOS stress (UV irradiation or H2O2)(# 24-3-11-5#33-3-11-5#34-3-17-5#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#20#3 clean-up -> XPcut(
# 27#28#14-5#3-30#2-9#24-3-17-5#2-3-17-5#20-#3-17-5#2-3-11-5-#3-#14#20-#28#3-#27#3-#29#35-#3-#14-5#3-30-#5#33-#3-17-5#34-3-17-5#36#37#3-17-5#2-3-11-5#2-3-11-5
September
'11/09/01(Thu)
- Colony PCR
- #34-3-17-5#3-14#3-29#35-#3-14-5#37#27#28#27#28#3-30 failed!,
# 14-5#3#20#3-14#3-29#3-14-5#3 Failed (Mixed with DMSO!),# 20#3-14#3-29#3-14-5#3-27#20-#3-17-5#36#3#20#14-5
- #34-3-17-5#3-14#3-29#35-#3-14-5#37#27#28#27#28#3-30 failed!,
'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#3-14#3-14-5#3-29#20-3-17-5#3-27#3#20#37#3-14#3-29#3-14-5#20-3-17-5#3-27 PCR products were OK but all purified products were lost.
**
# 20-3-17-5#3-27#3-14#3-14-5#3-29#20-3-17-5#3-27#3-14-5#3-29#3#20#37#3-14#3-29#3-14-5#20-3-17-5#3-27#3-14-5#3-29
- #20#3-14#3-14-5#3-29#20-3-17-5#3-27#3#20#37#3-14#3-29#3-14-5#20-3-17-5#3-27 PCR products were OK but all purified products were lost.
**
'11/09/03(Sat)
- Gel extraction
- Ligation & Transformation
- #20-#3-14-5#2-#3-14-5#3-27-#5#3-27-#5#37-#5#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#2-#3-14-5#3-27-#5#3-29-#5#37-#5#20-#28#36#2#5#20-3-14-5#20#20
'11/09/05(Mon)
- Colony Diffusion Assays (#2-9#20#20-3-14-5#20-3-17-5#2-#3-14-5#3-27-#5#3-29-#5#37-#5#20-#28#36 clean-up -> Digest(
# 3-29-#5#37-5#20-3-14-5#37-5#36 Asp -> all failed(.125%, .25% Agar) *Culture for Miniprep(# 2-3-14-5
'11/09/06(Tue)
- Gel extraction(#37-5#3-29-5#2-3-14-5#20-3-14-5#20-3-17-5#36#3-29-5#37-5 Purification -> Digest(
# 3-29-5#37-5#3-27-#5#3-30-#5#2-3-11-5-#5#2-9 4℃ storage *Ninhydrin sol. preparation (Not dissolvable!) -> 4℃ storage
'11/09/07(Wed)
- Colony PCR(#3-27-5#3-30-5#36#36#3-29-5#37-5#36#3-29-5#37-5#3-27-5#3-29-5#37-5#36 Digest(
# 3-14-5#3-29-5#37-5#3-27-5#36#2#2-3-17-5#3-14-5#34#2#34#36#3-27-5#3-14-5#2-3-17-5#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-#1-#3-17-5#33#34#36#1#2#20#3
'11/09/08(Thu)
- Making competent cell (WT), LB(amp) plate
- Frozen stocks(#36#3-14-5#3-27-5#33-3-14-5#34-3-14-5#36-3-14-5#20-3-29-5#2-3-11-5#2#34#36#2-3-17-5#3-27-5#3-14-5#35#31#3-14-5#3-27-5#3-29-5#37-5#2#34#36#2-3-17-5#3-14-5#34-3-14-5#36-3-14-5#20-3-29-5#20-3-14-5 Store at -20C(
# 33-3-11-5#34-3-14-5#20-3-29-5#37-5#35#31#31#3-#37-5#36-3-29-5#3-14-5#2-3-17-5#20-#3-27-5#34-3-17-5#36-3-14-5#20-3-29-5#2-3-11-5#34-3-14-5#36-3-14-5#20-3-14-5
'11/09/09(Fri)
- Making competent cell (WT), TB
- Colony PCR(#35-#31-#3-#37-5#36-#3-29-5#3-14-5-#2-3-17-5#20-3-27-5#3-14-5#3-17-5#31#3-37-5#36-3-29-5#3-14-5-2-3-17-5#3-11-5#3-14-5#3-17-5#31#3-37-5#36-3-29-5#34-3-14-5#36-3-14-5#20-3-29-5#2-3-11-5#34-3-14-5#36-3-14-5#20-3-14-5#3-11-5#3-11-5#29#36-3-14-5#2-3-17-5#20#3-27-5#2-3-11-5#20-3-29-5#20-3-29-5#31-#3-37-5#36-3-29-5#3-14-5-2-3-17-5 Luciferase Assay(
# 24-3-17-5#34-3-17-5 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-#3-37-5#36-3-29-5#3-14-5-2-3-17-5#3-14-5#3-17-5#31#3-37-5#36-3-29-5#36-3-14-5-#2-3-17-5#20#3-37-5
'11/09/11(Sun)
- Digest from Miniprep products
- #3-14-5-2-3-17-5#36-3-14-5#20 O.N.
*Colony PCR
**
# 3-11-5#29#20-#3-27-5#20-#3-37-5#36-#3-14-5-2-3-17-5#3-14-5-2-3-17-5#20-3-37-5#29#20#2-3-17-5#36-3-14-5-2-3-17-5#20#3-14-5-2-3-17-5#1#3-14-5-2-3-17-5 Digest for >1h -> Heat kill for 20min (# 20-3-37-5#36-3-14-5-2-3-17-51h -> Heat kill for 20min (# 36-3-14-5-2-3-17-5#2-3-17-5#20-3-14-5#2-3-17-5#36-3-14-5-2-3-17-5#20-3-37-5#36-3-14-5-2-3-17-5#20-3-37-5#20-3-29-5#20-3-29-5
- #3-14-5-2-3-17-5#36-3-14-5#20 O.N.
*Colony PCR
**
'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#36-3-14-5-2-3-17-5-#20-3-37-5-#20-3-37-5#2-3-11-5
Note:
# 36-3-14-5-2-3-17-5-#20-3-37-5-#20-3-37-5#36-3-14-5-2-3-17-5#20-3-37-5#20#29#2-3-17-5#2-3-37-5#36-3-14-5-2-3-17-5#20-3-37-5#20-3-37-5#2-9#20-3-29-5#20#36-3-14-5#2-3-17-5#36-3-14-5-2-3-17-5#20-3-37-5#20-3-37-5#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-#36-3-14-5-2-3-17-5#20-3-37-5#36-3-14-5-2-3-17-5#20-3-37-5#3-27-5#20#36-3-29-5#20-3-37-5
- #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#36-3-14-5-2-3-17-5-#20-3-37-5-#20-3-37-5#2-3-11-5
Note:
'11/9/13 (Tue)
- Colony PCR
- #36-3-14-5-2-3-17-5#20-3-37-5#36-3-14-5-2-3-17-5#20-3-37-5#3-27-5#20#36-3-29-5#20-3-37-5#36-3-14-5-2-3-17-5-20-3-37-5#20-3-14-5-2-3-17-5#1-3-14-5-2-3-17-5
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#20-3-37-5#20-3-29-5#2-9#20-3-29-5#1-3-14-5-2-3-17-5#20-3-14-5-2-3-17-5#36-3-14-5-2-3-17-5-20-3-37-5#2-3-11-5#20-3-37-5#20-3-29-5#3-27-5#31#20#3-29-5#20#3-27-5#3-14-5-2-3-17-5#1-3-14-5-2-3-17-5#20-3-14-5-2-3-17-5#20-3-37-5
- #36-3-14-5-2-3-17-5#20-3-37-5#36-3-14-5-2-3-17-5#20-3-37-5#3-27-5#20#36-3-29-5#20-3-37-5#36-3-14-5-2-3-17-5-20-3-37-5#20-3-14-5-2-3-17-5#1-3-14-5-2-3-17-5
Note: Seemes OD600 should NOT exceed 0.4~0.5, or intracellular luciferase will be saturated.
*Miniprep.
**
'11/09/14(Wed)
- Colony PCR
- #3-27-5#20#20-3-14-5-2-3-17-5#1-3-14-5-2-3-17-5#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#20#20#20-3-14-5-2-3-17-5#3-14-5-2-3-17-5#3-17-5#3-11-5#19#20#3-29-5#36-3-14-5-2-3-17-5#20-3-37-5#1-3-14-5-2-3-17-5#20-3-14-5-2-3-17-5#20-3-37-5#20-3-29-5#2-3-11-5#3-27-5#31
'11/09/15(Thu)
- Colony PCR(#19#36-3-14-5-2-3-17-5-20-3-37-5#20-3-29-5#20#20-3-14-5-2-3-17-5#3-14-5-2-3-17-5#3-17-5#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#20#3-11-5#20-3-29-5#19#3-14-5-2-3-17-5#3-14-5-2-3-17-5#36-3-14-5-2-3-17-5#20-3-37-5#3-27-5#20#3-17-5#3-14-5#3-14-5-2-3-17-5#20-3-14-5-2-3-17-5#20-3-14-5-2-3-17-5
'11/09/16(Fri)
- Colony PCR
- sulAp(ES)-#3-14-5-2-3-17-5#3-14-5-2-3-17-5#36-3-14-5-2-3-17-5#20-3-37-5#3-27-5#20#3-17-5#3-14-5#3-14-5-2-3-17-5#20-3-14-5-2-3-17-5#36-3-14-5-2-3-17-5#20-3-37-5#3-11-5#20-3-29-5#19#3-27-5#36-3-14-5-2-3-17-5#20-3-29-5#3-27-5#36-3-14-5-2-3-17-5#20-3-14-5-2-3-17-5
'11/09/17(Sat)
- Colony PCR (retry)
- #3-14-5#3-17-5#36-3-14-5-2-3-17-5-20-3-37-5#3-14-5#3-17-5#20-#3-27-5#20-3-29-5
'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#3-14-5#3-17-5#20-#3-27-5#3-14-5-2-3-17-5#31-
'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#36-3-14-5-2-3-17-5-20-3-37-5#33-3-14-5-2-3-17-5#24-#3-14-5-2-3-17-5#20-#3-27-5#36-3-14-5-2-3-17-5-20-3-37-5#33-3-14-5-2-3-17-5#20-3-29-5#1#20#36#33
'11/09/20(Tue)
- colony PCR(20-3-27-5, #24-3-14-5-2-3-17-5#3-27-5 Digest(
# 29#20-3-29-5#3-29-5#20#3-11-5#3-14-5#3-17-5#31#36-3-29-5#20-3-37-5
'11/09/21(Wed)
- Making plates
- Gel extraction(#20#29#3-29-5#20-3-29-5#20#3-27-5#24#3-14-5-2-3-17-5#29#3-29-5#20-3-29-5#3-14-5-2-3-17-5#36#33-3-14-5-2-3-17-5#3-14-5-2-3-17-5#3-14-5-2-3-17-5#3-29-5#36-3-14-5-2-3-17-5
'11/09/22(Thu)
- Ligation (retry)
- #3-27-5#20#3-14-5-2-3-17-5-#24#29#3-29-5#20-3-29-5#3-14-5-2-3-17-5#36culture for mini prep),
# 33-3-14-5-2-3-17-5Maybe the amplification was failed)) *Gel extraction(# 3-14-5-2-3-17-5#3-29-5#3-14-5-2-3-17-5#20-3-37-5#3-14-5-2-3-17-5#20-3-37-5#3-29-5#20-3-37-5
- #3-27-5#20#3-14-5-2-3-17-5-#24#29#3-29-5#20-3-29-5#3-14-5-2-3-17-5#36culture for mini prep),
'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#3-29-5-#20-3-29-5-#36#3-29-5#3-29-5-#20-3-29-5-
