Team:UT-Dallas/Notebook/8-22

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<h1 class="firstHeading" align="center">Friday, August 22, 2014</h1>
<h1 class="firstHeading" align="center">Friday, August 22, 2014</h1>
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<p class="tab"> Today is also long day. (We almost repeated everything we did yesterday: inoculation, miniprep, ligation, and transformation.</p>
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<p class="tab"> Today is also long day. (We almost repeated everything we did yesterday: inoculation, miniprep, ligation, and transformation. Also, sequencing results are here.</p>
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<p class="tab"> <b>Second batch (B2)</b>: We inoculated some toxT gRNA. Today we are minipreping it. (It already passed the test digest.</p>
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<p class="tab"> <b>Second batch (B2)</b>: </p>
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<p class="tab"> <b>Third batch (B3)</b>: Yesterday, we inoculated 2 reporter-Chlora colonies from each plates. Today we digest them, gel purify, ligate, and transform them onto Carb back bone, along with re-ligating and re-transforming ctxA that kept failing. Gel test for some were not good so we inoculated a few more (from tcpR).</p>
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<UL>
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<p class="tab"> <b>Forth batch (B4)</b>: We transformed B4 reporters on promoter-Chlora. The plates are good today. There's one colonies on the negative and the colonies on other plates appear with different sizes, so we are picking 2 colonies from each plates to inoculate.</p>
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<LI>toxT: got very high noise in the background. We are possibly re-sequencing them.</p>
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<p>We remade some gRNA from the primers. Unfortunately, yesterday we didn't have the machine for overnight ligation. So we will ligate it tonight.</p>
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<LI>tcpA: (gRNA+Promoter) first clone was wrong after sequencing. We have two clones so we will re-test-digesting-2%-gel them and send them out for sequencing.
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</UL>
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<p class="tab"> <b>Third batch (B3)</b>: Yesterday, we transformed reporter-Carb; today, we inoculate them. The colonies are not very big, we have about 3 colonies on the negative; however, the other plates are very good: they has at least twice-fold the colonies on the negative.</p>
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<p class="tab"> <b>Forth batch (B4)</b>: We inoculated yesterday. We are minipreping, digesting, ligating onto Carb back bone, and transforming them</p>
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<p>tcpQ is being re-digested and re-tested on 2% gel to make sure of our clone. We tested it on 1% gel but it created a smear and was dubious.</p>
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<p class="tab">Ran out of PstI-HF!!! Need to order!!!</p>
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<br>
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<p class="tab"> <b>Tomorrow:</b> For B3, we need to re-do whatever we did today because of new inoculation of the old plates. Meaningly, miniprep, digest, gel purify, ligate, and transform newly inoculated tcpR. we should have colonies of B3 reporters on the plates we are transforming today. We should miniprep the B4 reporters we inoculated today. For gRNA of B4, ligation will be done and we will transform them tomorrow. Saturday, we will inoculate. Sunday, we will miniprep. Next week, on Monday, we should have everything ready for test digest and then finally send the last batch of sequencing!</p>
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<p class="tab"> <b>Tomorrow:</b> For B4 reporters and B3 gRNA, inoculation. B3 reporters: miniprep. Sunday, we will miniprep. Next week, on Monday, we should have everything ready for test digest and then finally send the last batch of sequencing!</p>
    
    
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<LI> B3 reporter vectors on Chlora: Glycerol Stock + Miniprep + Digest + Gel extract and purify + ligate + transform
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<LI> B3 reporter vectors on Chlora: Inoculate
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<LI> Inoculate 2 more colonies from tcpR plates.
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<LI> B4 reporters-chlora: Glycerol stock + Miniprep + digest + gel extract + purify + ligate + transform onto carb back bone.
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<LI> B4 reporter vectors on Chlora: Inoculate 2 colonies from each plate acfA, acfB, tcpT, tcpQ
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<LI> Inoculate acfA(3)-chlora reporter.
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<LI> ctxA: ligate + transform
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<LI> Miniprep toxT gRNA.
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<LI> Overnight ligation of primers gRNA under promoter: acfA, acfB, tcpF
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Sam and Kortne during our down time in the lab
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      <img src="http://i.imgur.com/5xLPlXC.jpg" height="320px">&nbsp;<img src="http://i.imgur.com/7cZj3IZ.jpg" height="320px">
 
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      <br> Terrible, terrible gel! We punctured the gel! The lane bled over each other. We had to make another gel to purify. <br><b><i>(In order, i5.47.1-2-3-4-X-5-6-7-8-9-10-11-12)</i></b></center>
 
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      <br> Updating the wiki notebook from the slowest computer in the building because currently we can't log into any other computer. Rishika is gel-ing the ctx-s.
 
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Latest revision as of 14:31, 17 October 2014

Home Team Official Profile Project Parts Modeling Notebook Safety Attributions Human Practices

Friday, August 22, 2014

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Today is also long day. (We almost repeated everything we did yesterday: inoculation, miniprep, ligation, and transformation. Also, sequencing results are here.

Second batch (B2):

  • toxT: got very high noise in the background. We are possibly re-sequencing them.

  • tcpA: (gRNA+Promoter) first clone was wrong after sequencing. We have two clones so we will re-test-digesting-2%-gel them and send them out for sequencing.

Third batch (B3): Yesterday, we transformed reporter-Carb; today, we inoculate them. The colonies are not very big, we have about 3 colonies on the negative; however, the other plates are very good: they has at least twice-fold the colonies on the negative.

Forth batch (B4): We inoculated yesterday. We are minipreping, digesting, ligating onto Carb back bone, and transforming them

tcpQ is being re-digested and re-tested on 2% gel to make sure of our clone. We tested it on 1% gel but it created a smear and was dubious.


Ran out of PstI-HF!!! Need to order!!!


Tomorrow: For B4 reporters and B3 gRNA, inoculation. B3 reporters: miniprep. Sunday, we will miniprep. Next week, on Monday, we should have everything ready for test digest and then finally send the last batch of sequencing!
Today's tasks:
  • B3 reporter vectors on Chlora: Inoculate
  • B4 reporters-chlora: Glycerol stock + Miniprep + digest + gel extract + purify + ligate + transform onto carb back bone.
  • Inoculate acfA(3)-chlora reporter.
Sam and Kortne during our down time in the lab

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