Difference between revisions of "Team:Uppsala/Notebook"

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<h3>Week: 19/06- 22/06</h3>
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- Testing of the competent cells made last week was done with the competent cells test kit. There was growth suggesting that competent cells were working good.<br>
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- To test if the restriction digestion, ligation and transformation worked well, they were repeated again with the newly prepared and tested competent cells above. No growth was observed even when high concentrations were plated, suggesting that the fault was in the Restriction digestion, ligation steps<br>
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<h3>Week: 26/06-30/06</h3>
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- On expert suggestion, it was understood that the enzymes need few extra bases on the ends for efficient activity. Thus we designed and ordered primers that would add overhangs to our constructs. Other reason could be that the promoters were too strong and the genes were toxic to the cell. So a set of primers were also designed to remove promoter from between the prefix and RBS.<br>
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- To circumvent the high toxicity of the gene, we decided to clone the devices in low copy plasmids. For that low copy plasmids were identified from the master strain database of glycerol stock at the lab, grown overnight in agar plates, grown overnight in LB and plasmids were purified. <!--Concentration of the purified are as following--><br>
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<!--<table>
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  <tr>
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    <th>Colony (database nr.sample nr)</th>
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    <th>Concentraton (ng/μL)</th>
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    <th>Avg. Conc. (ng/μL)</th>
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    <th>Purity (260/280 nm)</th>
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  </tr>
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  <tr>
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    <td>c11.1</td>
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    <td>35.3; 33.9</td>
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    <td>34.75</td>
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    <td>1.86; 1.86</td>
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  </tr>
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  <tr>
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    <td>c11.2</td>
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    <td>34.0; 34.2</td>
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    <td>34.1</td>
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    <td>1.76; 1.82</td>
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  </tr>
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  <tr>
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    <td>c11.3</td>
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    <td>29.2; 29.1;</td>
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    <td>29.15</td>
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    <td>1.81; 1.82</td>
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  </tr>
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  <tr>
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    <td>c15.1</td>
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    <td>32.4; 31.2</td>
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    <td>31.8</td>
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    <td>1.91; 1.88</td>
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  </tr>
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  <tr>
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    <td>c15.2</td>
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    <td>40.3; 39.9</td>
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    <td>40.1</td>
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    <td>1.90; 1.87</td>
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  </tr>
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  <tr>
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    <td>c15.3</td>
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    <td>36.5; 36.9</td>
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    <td>36.7</td>
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    <td>1.84; 1.88</td>
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  </tr>
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  <tr>
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    <td>c58.1</td>
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    <td>29.8; 30.1</td>
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    <td>29.95</td>
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    <td>1.94; 1.93</td>
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  </tr>
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  <tr>
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    <td>c58.2</td>
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    <td>29.4; 28.7</td>
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    <td>29.05</td>
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    <td>1.87; 1.90</td>
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  </tr>
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  <tr>
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    <td>c58.3</td>
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    <td>36.7; 27.2</td>
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    <td>31.95</td>
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    <td>1.71; 1.89</td>
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  </tr>
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  <tr>
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    <td>c98.1</td>
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    <td>38.0; 38.6</td>
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    <td>38.3</td>
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    <td>1.91; 1.86</td>
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  </tr>
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  <tr>
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    <td>c98.2</td>
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    <td>38.9; 37.5</td>
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    <td>38.2</td>
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    <td>1.91; 1.93</td>
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  </tr>
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  <tr>
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    <td>c98.3</td>
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    <td>29.6; 29.6</td>
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    <td>29.6</td>
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    <td>1.90; 1.91</td>
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  </tr>
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</table><br>-->
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- Digestion was tested with X and S to optimize the protocol for digestion<br>
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Revision as of 14:32, 31 October 2017