Team:Alberta/Notebook

From 2012.igem.org

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<font size=5> &nbsp;&nbsp;&nbsp;iGem Diary </font>
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<font size=5> &nbsp;&nbsp;&nbsp;iGem Notebook</font>
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Rick and Tom recruited and learning basic laboratory protocols, such as making competent cells, restriction digests, cell transformation, gel electrophoresis.
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Rick and Tom began coming in to the lab regularly and started learning basic laboratory protocols, such as making competent cells, restriction digests, cell transformation, gel electrophoresis, ect.
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Today Tom finally managed to get a PCR to work, though it took him about twelve attempts.
Today Tom finally managed to get a PCR to work, though it took him about twelve attempts.
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Competent cells were made today using a standard procedure which took the entire day.
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Competent cells were made today using a standard procedure which took the entire day. The competent cell procedure is described elsewhere in this wiki.
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The competent cells were tested with basic puc19 transformation, and the transformation worked
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The competent cells were tested with basic puc19 transformation, and the transformation worked with acceptable efficiency.
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Today multiple PCRs were run on puc19 with two of the color genes and a C1 gene.
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Today multiple PCRs were run on puc19 with two of the color genes and a repressor gene.
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Made pUC19 plasmid with the new color genes and C1 and transformed it into cells.
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Made pUC19 plasmid with various color genes and C1 and transformed it into cells.
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We cloned a variety of different (9) promoters into our color gene plasmids, in order to get a sense of relative promoter strengths. we also seqencing those construct to check they are correct.
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We cloned a variety of different promoters (nine total) into our color gene plasmids in order to get a sense of relative promoter strengths. We also sent off to sequencing those construct to check that they are correct.
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We PCR new RBS  colour genes and regulatory promoters. we then made those constructs into kanamycin resistant plasmid backbone and transform them into Top 10 cells.
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We used PCR to add new RBS  colour genes and regulatory promoters to our constructs. We then pieced those constructs into kanamycin resistant plasmid backbone and transform them into Top 10 cells.
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Torrin and Sarah Join. we perform the same work again on TG1.  
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Torrin and Sarah join the team. We performed the same work on the TG1 strain of E.Coli as a learning exercise.  
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Tom checked overnights of origin cut site strains and chose colonies to be sequenced.
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Creation of a plasmid with a cut site near the origin of replication began today. This plasmid will be used in order to modify the promoter on the plasmid's origin of replication.
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Today Tom Used the successful origin cut site plasmid with Amp Resistance and replaced that resistance with Chlr.
Today Tom Used the successful origin cut site plasmid with Amp Resistance and replaced that resistance with Chlr.
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Torrin and Sarah are creating and testing chemical gradient plates, and learning additional laboratory procedure.
 
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Rick and Spencer is making competent cells of TG1 so that we can test our current parts in an E.Coli strain that grows faster than our current Top10.
 
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Rick and Easwar are drop in LacI and Tet R repressor into chloranphenicol construct.
 
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July 11
 
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We continued our experiments with making chemical gradients on agar plates.
 
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July 12-17
 
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we successfully get the LacI and TetR repressors into chlr contstruct and co-transfrom with RTa and RL we created from july 7. the result showed seemed it is working and we have white colony.Then we decide to send TetR and LacI construct for sequencing. The result showed LacI and TetR sequence were good.
 
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July 18
 
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The Lac and Tet promoter pieces have been successfully added to plasmids via specially designed PCR primers, and will control the origin of replication on said plasmids. We will start testing them over the next couple of weeks.
 
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July 19
 
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we decide to send TetR and LacI construct for sequencing. The result showed LacI and TetR sequence were good.
 
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July 20-25
 
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co-transformation of lacI/tetR promoter with red gene and repressor (lacI and TetR) gene were successfully, however, when we do the reverse experiment (LacI/tetR with non-repressor gene) with isolated white colonies were not many red colonies.
 
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<div class="underline"><font size=5>Augest</font></div>
 
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July 30- Aug 13
 
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Tom on vancation. Rick take over Tom’s work continuing on the research on the process of controlling plasmid copy number in the cell. we redesign the plasmid so that Tom’s original desing wont killing cells by introducing lac operator or tet operator into the wild ori promotor with NsiI site beside it.
 
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Latest revision as of 21:56, 18 September 2012

   Team Alberta




   iGem Notebook


May


May.9-11.2012

Rick and Tom began coming in to the lab regularly and started learning basic laboratory protocols, such as making competent cells, restriction digests, cell transformation, gel electrophoresis, ect.


May. 24

Rick, Tom, Spencer, and Easwar learned how to use PCR today, with varying degrees of success.


May 31

Today Tom finally managed to get a PCR to work, though it took him about twelve attempts.

[Top page]


June


June 4

Made working stock of puc19 and then transformed Top10 with puc19. Overnight cultures were also set up in preparation for the making of competent cells


June 5

Competent cells were made today using a standard procedure which took the entire day. The competent cell procedure is described elsewhere in this wiki.


June 6

The competent cells were tested with basic puc19 transformation, and the transformation worked with acceptable efficiency.


June 7-8

Today multiple PCRs were run on puc19 with two of the color genes and a repressor gene.


June 11

Made pUC19 plasmid with various color genes and C1 and transformed it into cells.


June 18-21

We cloned a variety of different promoters (nine total) into our color gene plasmids in order to get a sense of relative promoter strengths. We also sent off to sequencing those construct to check that they are correct.


June 22- July 3

We used PCR to add new RBS colour genes and regulatory promoters to our constructs. We then pieced those constructs into kanamycin resistant plasmid backbone and transform them into Top 10 cells.

[Top page]


July


July 4-5

Torrin and Sarah join the team. We performed the same work on the TG1 strain of E.Coli as a learning exercise.


July 6

Creation of a plasmid with a cut site near the origin of replication began today. This plasmid will be used in order to modify the promoter on the plasmid's origin of replication.


July 9-10

Today Tom Used the successful origin cut site plasmid with Amp Resistance and replaced that resistance with Chlr.

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