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href="https://2011.igem.org/Team:ZJU-China/Notebook.html">July</a> <a
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href="https://2011.igem.org/Team:ZJU-China/August.html">August<br />
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</a> <a href="https://2011.igem.org/Team:ZJU-China/September.html">September<br />
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<p></p>
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<h1>Lab Notes - July</h1>
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<!--importantcontainer-->
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<h1>Biobrick Group</h1>
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<div id="page1">
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<h1>Week1</h1>
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<table id="notesheet"  border="1" cellspacing="0"
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cellpadding="1" style="vertical-align: middle">
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<tr>
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<td width="76"><strong>Day</strong></td>
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<td width="349"><strong>Note</strong></td>
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-
</tr>
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<tr>
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<td id="sheetleft">Jul.4th Monday</td>
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<td>
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-
<table id="intable" width="328" border="0" cellspacing="0"
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cellpadding="1">
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<td width="128">▪ Aerobic cultivation of DH5α</td>
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<td width="196">▪ Preparation of apparatus for the formation of
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biofilm</td>
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-
 
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</table>
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</td>
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</tr>
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<tr>
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<td id="sheetleft">Jul 5th Tuesday</td>
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<td>&nbsp;</td>
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</tr>
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<tr>
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<td id="sheetleft">Jul.6th Wednesday</td>
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<td>
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<table width="217" border="0" cellspacing="0" cellpadding="1">
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<tr>
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<td width="215">▪ Receiving primers ordered previously</td>
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-
 
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</tr>
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-
</table>
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-
</td>
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</tr>
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-
<tr>
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<td id="sheetleft">Jul.7th Thursday</td>
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<td>
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<table width="238" border="0" cellspacing="0" cellpadding="1">
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<tr>
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<td width="200">▪ Preparation of the aliquot of the primers</td>
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<td width="200">▪ Something wrong with a shaking incubator</td>
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-
</tr>
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-
</table>
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</td>
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</tr>
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<tr>
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<td id="sheetleft">Jul.8th Friday</td>
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<td>
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<table width="222" border="0" cellspacing="0" cellpadding="1">
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<tr>
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<td width="155">▪ Preparation of culture plates for the
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transformations</td>
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</tr>
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</table>
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</td>
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</tr>
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<tr>
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<td id="sheetleft">Jul.9th Saturday</td>
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<td>
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<table width="313" border="0" cellspacing="0" cellpadding="1">
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<tr>
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<td width="127">▪ Preparation of culture plates for the
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transformations</td>
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<td width="182">▪ protocols of transformation</td>
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-
</tr>
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</table>
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</td>
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</tr>
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<tr>
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<td id="sheetleft">Jul.10th Sunday</td>
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-
<td>
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<table width="246" border="0" cellspacing="0" cellpadding="1">
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<tr>
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<td>▪ Several colonies were picked up and cultivated in 5mL LB
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medium. ▪ryosectioning of biofilm</td>
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-
</tr>
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-
</table>
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-
</td>
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-
</tr>
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-
 
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-
</table>
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-
</p>
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-
<h1>Week2</h1>
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-
<table id="notesheet" width="650" border="1" cellspacing="0"
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-
cellpadding="1">
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-
<tr>
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-
<td width="76"><strong>Day</strong></td>
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-
<td width="349"><strong>Note</strong></td>
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-
</tr>
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-
 
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-
<tr>
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-
<td id="sheetleft">Jul.11th Monday</td>
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-
<td>
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-
<table id="intable" width="541" border="0" cellspacing="0"
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-
cellpadding="1">
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<td width="128">▪ Set up new LB culture plates with ampicillin
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and kanamycin
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-
</p>
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-
</td>
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-
 
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-
 
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-
<td width="196">▪ Sterilization of Glycerol and <br />
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Preparation of 25mg/mL kanamycin</td>
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-
<td>▪Transformation of the parts mentioned on Jul.9th for the
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-
second time</td>
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-
<td>▪Observation the sections</td>
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-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul 12th Tuesday</td>
+
-
<td>
+
-
<table id="intable" width="560" border="0" cellspacing="0"
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-
cellpadding="1">
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-
 
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-
<td width="128">▪ Pick two colonies of each parts and cultivate
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-
them in LB medium</td>
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-
 
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-
 
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-
<td width="203">▪ Transformation of three parts(20J,20H.22B from
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Kit plates of 2011 Distribution ) which are related to the
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-
degradation of Cellulose</td>
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-
<td width="91">▪Min prep to isolate 10I,12I and 22M</td>
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-
<td width="130">▪Conservation of 10I,12I,22M and 11P</td>
+
-
</table>
+
-
</td>
+
-
</tr>
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-
<tr>
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-
<td id="sheetleft">Jul.13th Wednesday</td>
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-
<td>
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-
<table width="618" border="0" cellspacing="0" cellpadding="1">
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-
<tr>
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-
<td width="104">▪ Place the culture plate of 20J,20H and 22B in
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the fridge.</td>
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-
<td width="102">▪min prep to isolate 13K ▪onservation of 13K</td>
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-
<td width="163">▪estriction digest of the
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parts(12I,10I,22M,13K) with EcoRI and PstI</td>
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-
<td width="111">▪Gel electrophoresis to analyse restriction
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-
fragments</td>
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<td width="128">▪Test the Tm of Primers CP1&amp;CS,NP&amp;NS
+
-
with 13K. The result of Gel electrophoresis shows that 60.2℃ is the
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Tm of NS and NP, and 57.4℃ is the Tm of CS and CP1.</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.14th Thursday</td>
+
-
<td>
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-
<table width="618" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
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-
<td>▪ Test the Tm of the YFP,RFP,tetR,Vgb and nirB primers.
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-
The result of Gel electrophoresis shows that the Tm for the primers
+
-
of Vgb is 54℃ and the Tm for the primers of nirB is 55℃ The RCR of
+
-
YFP,RFP and tetR failed.</td>
+
-
 
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.15th Friday</td>
+
-
<td>
+
-
<table width="600" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
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-
<td width="155">▪ PCR(Phusion)<br />
+
-
▪ Digest 10I, 22M, 13K ▪ Used wrong cutter, digestion again.</td>
+
-
<td>▪ Run the results of PCR and the first digestion. The
+
-
annealing temperature of YFP needed change. The digestion results
+
-
confirmed</td>
+
-
<td>▪ Run the digestion results of second time. The bands are
+
-
confirmed.</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.16th Saturday</td>
+
-
<td>
+
-
<table width="620" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="127">▪ Cut the linearized pSB1k3 with E+P</td>
+
-
<td width="123">▪ Purify the digestion results of 22M, 13K, 10I</td>
+
-
<td width="133">▪ Confirm digestion of pSB1k3 by
+
-
electrophoresis, then purification</td>
+
-
<td width="113">▪ Test Tm of YFP<br />
+
-
▪ ligation: 22M+10I, 13K+10I</td>
+
-
<td width="114">▪ Tm of YFP is 54 degree ▪ transform the
+
-
ligation results.</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.17th Sunday</td>
+
-
<td>
+
-
<table width="620" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td>▪ PCR 22M<br />
+
-
▪ purify Vgb, RFP, RFP-tetR, YFP-tetR. Gibson PCR.</td>
+
-
<td>▪ ligation the purified the fragments in yesterday.</td>
+
-
<td>▪ 22M PCR<br />
+
-
▪ Transform the ligation results.</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
 
+
-
</table>
+
-
 
+
-
<p>&nbsp;</p>
+
-
<h1>Week3</h1>
+
-
<table id="notesheet" width="713" border="1" cellspacing="0"
+
-
cellpadding="1">
+
-
<tr>
+
-
<td width="87"><strong>Day</strong></td>
+
-
<td width="616"><strong>Note</strong></td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.18th Monday</td>
+
-
<td>
+
-
<table id="intable" width="541" border="0" cellspacing="0"
+
-
cellpadding="1">
+
-
 
+
-
<td width="128">▪ Genome extraction of E.coli</td>
+
-
 
+
-
 
+
-
<td width="196">▪ PCR of NirB<br />
+
-
▪ Cut 10I with X+P, cut 22M, 13K with E+S, cut pSB1k3 with E+P</td>
+
-
<td>▪ Ligate: 10I+13K, 10I+22M <br />
+
-
▪ Repeat NirB PCR</td>
+
-
<td>
+
-
<p align="left">▪ Culture 11P<br />
+
-
▪ Miniprep 10I, 22M, 13K</p>
+
-
</td>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul 19th Tuesday</td>
+
-
<td>
+
-
<table id="intable" width="615" border="0" cellspacing="0"
+
-
cellpadding="1">
+
-
 
+
-
<td width="140">▪ mini-prep: 10I+13K, 10I+22M</td>
+
-
 
+
-
 
+
-
<td width="213">▪Insert 10I+13K, 10I+22M into pSB1k3</td>
+
-
<td width="110">▪Transform: 5E, 3C, 7C, 1K, 1I from the
+
-
distribution plate</td>
+
-
<td width="144">▪Transform 10I+22M, 10I+13K</td>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.20th Wednesday</td>
+
-
<td>
+
-
<table width="610" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="170">▪Colonies of 22M+10I, 13K+10I, 5E, 3C, 7C, 1K,
+
-
1I confirmed</td>
+
-
<td width="102">▪ Gibson PCR</td>
+
-
<td width="250">▪Colony PCR 22M+10I, 13K+10I, 5E, 3C, 7C, 1K,
+
-
1I</td>
+
-
<td width="80">▪PCR NirB</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.21th Thursday</td>
+
-
<td>
+
-
<table width="618" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td>▪PCR verification of 22M+10I, 13K+10I, 5E, 3C, 7C, 1K, 1I
+
-
</td>
+
-
<td>▪ Gibson PCR ▪ Run the results of PCR verification. Bands
+
-
confirmed.</td>
+
-
<td>▪ Culture the 22M+10I, 13K+10I, 5E, 3C, 7C, 1K, 1I ▪
+
-
Purification of Gibson PCR results</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.22th Friday</td>
+
-
<td>
+
-
<table width="618" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="117">▪ PCR amplification of Gibson assembly results
+
-
</td>
+
-
<td width="144">▪ Mini prep: 22M+10I, 22.Presever in -20<br />
+
-
▪ Mini prep: 1K,1I,3C,5E,7C<br />
+
-
▪ Gibson Assembly fail.</td>
+
-
<td width="164">▪ PCR NirB by Phusion<br />
+
-
▪Repeat PCR by changing Pnibr to Gnirbr<br />
+
-
▪Cut the mini and medi prep results with E</td>
+
-
<td width="185">▪ Run the results of PCR and digestion. Fail in
+
-
PCR of NirB, succeed in 10I+3K and 10I+22M ligation.</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.23th Saturday</td>
+
-
<td>
+
-
<table width="613" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="237">▪Double digestion of 13K+10I, 22M+10I, pSB1c3
+
-
 
+
-
</td>
+
-
<td width="169">▪ Fail in purification of Medi prep</td>
+
-
<td width="201">▪ PCR NirB<br />
+
-
▪ Ligate NirB+13K+10I, Vgb+22M+10I</td>
+
-
 
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.24th Sunday</td>
+
-
<td>
+
-
<table width="608" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="244">▪ Gibson assembly: NirB+RFP+tetR<br />
+
-
▪ Cut results of Gibson assembly and pSB1c3.</td>
+
-
<td width="164">▪ Purify the ligation results in yesterday</td>
+
-
<td width="194">▪ ligate with backbone ▪ Culture 1I, 1K, 3C,
+
-
5E, 7C, 11P</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
 
+
-
</table>
+
-
<p>&nbsp;</p>
+
-
<h1>Week4</h1>
+
-
<table id="notesheet" width="691" border="1" cellspacing="0"
+
-
cellpadding="1">
+
-
<tr>
+
-
<td width="63"><strong>Day</strong></td>
+
-
<td width="630"><strong>Note</strong></td>
+
-
</tr>
+
-
 
+
-
<tr>
+
-
<td id="sheetleft">Jul.25th Monday</td>
+
-
<td>
+
-
<table id="intable" width="541" border="0" cellspacing="0"
+
-
cellpadding="1">
+
-
 
+
-
<td width="128">▪ Transform Pnirb+13k+10I+pSBK3-2</td>
+
-
 
+
-
 
+
-
<td>▪ Place the culture in 4 degree</td>
+
-
<td>▪ Medi-and mini-prep of five cultre</td>
+
-
<td>▪Gibson Assembly confirmation by gel</td>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul. 26th Tuesday</td>
+
-
<td>
+
-
<table id="intable" width="610" border="0" cellspacing="0"
+
-
cellpadding="1">
+
-
 
+
-
<td width="134">▪ Cut Pnirb, Pvgb,(E+S)<br />
+
-
▪ Cut 10I+13K, 10I+22M.(X+P)</td>
+
-
 
+
-
 
+
-
<td width="167">▪ Ligation: vgb+10I+22M, nirB+10I+13K,</td>
+
-
<td width="172">▪ Ligation: pSBK13+vgb+10I+22M,
+
-
pSBK13+nirB+10I+13K</td>
+
-
<td width="129">▪ Transform the ligation results.</td>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.27th Wednesday</td>
+
-
<td>
+
-
<table width="608" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="170">▪ No colony on the plate<br />
+
-
▪ Gibson assembly</td>
+
-
<td width="184">▪ Run the results of Gibson assembly</td>
+
-
<td width="248">▪ Excise the bands, the purify the DNA<br />
+
-
▪ Gibson Assenmly</td>
+
-
 
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.28th Thursday</td>
+
-
<td>
+
-
<table width="609" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="607">▪ 2011 China Meet-up @ Hefei</td>
+
-
 
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.29th Friday</td>
+
-
<td>
+
-
<table width="607" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td>▪ 2011 China Meet-up @ Hefei</td>
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.30th Saturday</td>
+
-
<td>
+
-
<table width="620" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
 
+
-
 
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
<tr>
+
-
<td id="sheetleft">Jul.31th Sunday</td>
+
-
<td>
+
-
<table width="600" border="0" cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td width="368">▪ Culture 22M+10I, 13K+10I</td>
+
-
<td width="228">▪ Cut a0H, 20J, 22B</td>
+
-
 
+
-
</tr>
+
-
</table>
+
-
</td>
+
-
</tr>
+
-
 
+
-
</table>
+
-
<br />
+
-
<br />
+
-
</div>
+
-
<!--page1-->
+
-
<div id="importantcontainer">
+
-
<div class="frame" id="important">
+
-
<div class="bgcolors" id="round">
+
-
<h1>Data/Protocol</h1>
+
-
</div>
+
-
</div>
+
-
</div>
+
-
<div id="framecontent">
+
-
<h1>Jul.13th Wednesday</h1>
+
-
 
+
-
<p><strong>Systems of restriction digestion with EcoRI and
+
-
PstI</strong></p>
+
-
<p>
+
-
<table style="margin: 15px;" width="110" height="200" border="1"
+
-
cellspacing="0" cellpadding="1">
+
-
<tr>
+
-
<td>Plasmid</td>
+
-
<td>1μL (>100ng)</td>
+
-
</tr>
+
-
<tr>
+
-
<td>EcoRI</td>
+
-
<td>1μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>PstI</td>
+
-
<td>1μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>10×Buffer Tango</td>
+
-
<td>2μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>ddH2O</td>
+
-
<td>15μL</td>
+
-
</tr>
+
-
</table>
+
-
<img src="https://static.igem.org/mediawiki/2011/c/c3/0713program.png"
+
-
width="200" height="200" /></p>
+
-
<p><strong>Temperature grad</strong><br />
+
-
56℃ 57.4℃ 60.2℃ 62.9℃ 64.3℃ 65.8℃ /p>
+
-
<div id="framecontent">
+
-
<p>&nbsp;</p>
+
-
 
+
-
<p><strong>PCR system (test the Tm of the primers
+
-
CP1&amp;CS, NP&amp;NS)</strong></p>
+
-
 
+
-
<table style="margin: 15px;" width="110" border="1" cellspacing="0"
+
-
cellpadding="1">
+
-
<tr>
+
-
<td>10×Buffer</td>
+
-
<td>2μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>dNTPs</td>
+
-
<td>0.5μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>primers CP1 (NP)</td>
+
-
<td>1μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>primers CS (NS)</td>
+
-
<td>1μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>Template</td>
+
-
<td>1μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>rTaq</td>
+
-
<td>0.2μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>H2O</td>
+
-
<td>14.5μL</td>
+
-
</tr>
+
-
<tr>
+
-
<td>Total</td>
+
-
<td>20μL</td>
+
-
</tr>
+
-
</table>
+
-
</p>
+
-
<p>Within each compartment are<strong> components</strong>: include
+
-
different types of biomass ,substrates , products. biomass is often
+
-
divided into active microbial species, inert cells, and extracellular
+
-
polymeric substances(EPS).</p>
+
-
<p>The components can undergo transformation, transport, and
+
-
transfer <strong>processes</strong>. For example, substrate is consumed,
+
-
and this leads to the synthesis of new active biomass.</p>
+
-
<p>All process affecting each component in each compartment are
+
-
mathematically linked together into a<strong> mass balance
+
-
equation</strong> that contains rate terms and parameters for each process.</p>
+
-
<p><strong>Model Selection:</strong>Many kinds of Mathematics models
+
-
have been founded to describe a system of biofilm. Models of different
+
-
dimensions (1d, 2d, 3d) focus on different properties of a biofilm.
+
-
Since we care most about the oxygen concentration gradients
+
-
perpendicular to the substratum, <strong>numerical
+
-
1-dimensional dynamic model(N1)</strong> would be a proper choice for us.</p>
+
-
</div>
+
-
<!--bgcolors--></div>
+
-
<!--important--></div>
+
-
<!--importantcontainer-->
+
-
<p style="clear: both;"></p>
+
-
<div id="page2">
+
-
<div class="block" id="nsheet"><img
+
-
src="http://ung.igem.org/wiki/images/4/42/Biofilmcomp.png" width="640"
+
-
height="464" style="margin: 5px;" />
+
-
<p>&nbsp;</p>
+
-
</div>
+
-
<div class="block" id="nsheet">
+
-
<h1>Early July</h1>
+
-
<p>Pre-experiments with biofilm formation with circular and non
+
-
circular silicone tube, 24 well plate on different support including
+
-
glass, paper, plastic film, rubber. The final material are used based on
+
-
the easiness biofilm form on them and on the easiness to observe under
+
-
microscope.</p>
+
-
<img
+
-
src="https://static.igem.org/mediawiki/2011/b/be/20110711-3nP4100X0877.jpg"
+
-
width="400" style="margin-left: 20px;">
+
-
<p>&nbsp;</p></div>
+
-
<div class="block" id="nsheet">
+
-
<h1>28th July</h1>
+
-
<p>13.30: DH5α 11p 5mlX4<br />
+
-
23.00: silicone tube set at 37℃ /p>
+
-
</div>
+
-
<div class="block" id="nsheet">
+
-
<h1>29th July</h1>
+
-
<p>13.00: LB culture 50ml with circular culture medium. LB culture
+
-
50ml with noncircular culture medium.</p>
+
-
 
+
-
</div>
+
-
<div class="block" id="nsheet">
+
-
<h1>31st July</h1>
+
-
<p>13.00: -80℃ storing silicone tube. A thick white and loosely bond
+
-
substance is seen on the inner wall of the 5mm silicone tube, and on the
+
-
inner wall of the 1mm silicone tube a flatter and smoother white
+
-
substance. Especially obvious where the tube turns. Possibly because the
+
-
speed of culture flow is slower.</p>
+
-
 
+
-
</div>
+
-
</div>
+
-
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<p><a name="jumptop" id="bb">&nbsp; Biobrick Group&nbsp;</a>|<a
+
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Revision as of 17:36, 19 October 2011

  1. REDIRECT ZJU-China/Notebook
Retrieved from "http://2011.igem.org/Team:ZJU-China/Notebook.html"