SBB11Ntbk-Helen Shi

Helen Shi 15:49, 18 February 2011 (EST)

Today, I performed a gel cleanup for my digested part, jtk2936, for which I ran a preparation gel yesterday. I now have 8.5 uL of jtk2936 DNA. Next class, I will digest this part, ligate it with my plasmid vector, and transform it into E. coli.

For my other part, sbb1136, I should also digest it next class before ligation and transformation.

Helen Shi 13:38, 17 February 2011 (EST)

Today, I got my PCR back and then I ran an analytical gel to check that the size of my PCR product is 531 basepairs. From the gel picture, my PCR product appears to be about 500 basepairs, which is correct. Thus, I continued on to the purification of my PCR product with a Zymo Cleanup; I eluted 33 uL of my sbb1136 part. Tomorrow, I will digest my PCR product with EcoRI and BamHI.

021711-AnalGel1, Last column (~500 bp)

I also digested the plasmid pBjk2741-jtk2936 with EcoRI and BamHI restriction enzymes to cut out the HlyE part. After an hour of digestion, I am performing a preparation gel to extract the digested HlyE part (1095 bp). Tomorrow, I may ligate and transform my HlyE part.

021711-PrepGel2, Column 3 (~1000 bp)

Helen Shi 13:44, 15 February 2011 (EST)

Today, I set up the PCR for my sbb1136 part using the prediluted oligos ss63r and ss63f (10 uM). Since the template for my part came from the MG1655 genome, I used the Expand buffer and polymerase for my PCR. I set up my PCR reaction with 1 uL each of my 10 uM oligos. From my construction file, my predicted PCR product has a size of 531 bp so the PCR file to be used is 2K55.

Helen Shi 04:29, 8 February 2011 (EST)

Hi!