Difference between revisions of "Team:UCLA/Notebook/Spider Silk Genetics/4 May 2015"
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class="td11">2CA</td><td style="white-space:nowrap; text-align:left;" id="TBL-4-4-2" | class="td11">2CA</td><td style="white-space:nowrap; text-align:left;" id="TBL-4-4-2" | ||
class="td11">11.38 </td><td style="white-space:nowrap; text-align:left;" id="TBL-4-4-3" | class="td11">11.38 </td><td style="white-space:nowrap; text-align:left;" id="TBL-4-4-3" | ||
− | class="td11"> | + | class="td11">2.33 </td></tr></table></div> |
<br /> <div class="caption" | <br /> <div class="caption" | ||
><span class="id">Table 1: </span><span | ><span class="id">Table 1: </span><span |
Latest revision as of 04:22, 17 May 2015
5/4/2015
1 BsaI Digestion Test for 2AB
Tested digestion of 5 ug of plasmid to see efficiency of digestion.
Amount (uL) | |
BsaI | 2 uL |
2AB (287 ng/uL) | 14.7 uL |
Cutsmart | 5 uL |
ddH2O | 28.28 |
Total | 50 uL |
50 C | 2 hr |
65 C | 20 min |
Results are shown below:
∙ The digestion process works, so we’ll be proceeding with this protocol.
∙ We gel extracted the digested insert band. The end result was 11.9 ng/uL, A (260/280): 4.4
2 BsaI Digestion for Post-elution primer amplified MaSp
We BsaI digested the PCR products from previous PCRs using the following protocol
2.1 BsaI Digest Protocol
2AB(61.43 ng/uL) | 2BC* | CA (56.65 ng/uL) | |
BsaI | 2 uL | 2 uL | 2 uL |
DNA (500 ng) | 8.14 uL | 18.01 uL | 8.83 uL |
10x Cutsmart | 5 uL | 5 uL | 5 uL |
ddH2O | 34.86 uL | 25 uL | 34.17 uL |
Total | 50 uL | 50 uL | 50 uL |
50 C | 2 hr | ||
65 C | 20 min |
*For 2BC, we combined 8.84 uL of 33.9 ng/uL solution and 9.17 uL and 33.9 ng/uL solution to get 500 ng total.
2.2 Purification
∙ PCR purified the post-elution PCR digestion products and got the following yields:
∙ Yield and absorbance for the digested inserts are not very good. This might be a problem due to the PCR process, even though product was generated.
3 Bacterial Propagation
∙ Grew 7 mL each of AB, BC, CA from glycerol stock for DNA extraction tomorrow.