Tuesday June 22, 2010

Patrick: Being the highly intelligent person that I was, I realized that yesterday I had constructed the R0040 onto the original E0430, which was on a pSB1A2 plasmid. I plated on kanamycin agar plates. Thus, no growth. I will retry the construction once more using the plasmid-switched E0430. We also did a miniprep of E0420.

I also began the screening of the cpxP promoter. When we had designed the primers to PCR out the promoter from genomic DNA, we had added XbaI and SpeI sites to the primers, which are non-directional and may scar. Thus, we will try and digest 10 trials of cpxP over the next few days with these two enzymes, and hopefully pick out those that do separate into separate bands in a gel.

But first, the cpxP promoter needs to be placed in a plasmid for future BioBrick usage. This will be done tomorrow.

Dev: Today, I did a construction of E0040 + B0015. I inserted E0040 into the AK plasmid of B0015. E0040 was with EcoRI and B0015 was cut with EcoRI and XbaI. The vector was Antarctic phosphatased to prevent self ligation. The construct was transformed into competent cells and plated.