Tuesday June 15, 2010

Dev, Jeremy, and Chris: Today, we spent the day redoing transformations and constructions that failed to work from yesterday. We observed the plates that we grew with the constructions of J13002-E0032, J13002-E0040, R0040-J23032, and B0015-R0040. There was no growth observed on any of the plates and thus, they had to be redone. The registry parts I0500, K239000, K274210, and K135000 were observed. There was growth on all the streak plates except for the second version of I0500 which showed no growth once again. We ran out of the original I0500 arabinose inducible promoter and it showed inconsistent growth from the Registry tests. The new one was sent from a different location and once again, showed no growth. Today, we restreaked I0500 and attempted to innoculate it directly into overnight cultures. We also made overnight cultures for many of the parts that have been transformed so far from the Registry including: R0040, E0040, E0032 etc.

Emily: Today I did a Colony PCR as a final verification of my I0500-B0034 consruct, using BBK-CP primers. If it looks good, I will send a colony down for sequencing tomorrow.

Himika: Today, I checked the colonies that were grown from the construction of E1010-B0015. No colony growth were observed. From the trouble that our entire team has been having with construction and subsequent transformation, we concluded that our enzymes were possibly faulty or the ratios that we have been using are faulty. I decided to give it one more try with the restrictrion digest, ligation, and subsequent transformation to Top10 competent cells.

Alex, Patrick, Raida: We received some primers that can be used to PCR the CpxP promoter system from genomic DNA, which we intend to use to detect cellular stress and protein misfolding. Thus, we did a colony PCR of a colony from our plate stock using these primers. This would be a colony from the streak plate of the R0040-E0430 construct from June 14th.

We also did a restreak and overnight culture of the following: two K239000 plates, two E0430 plates, and two E0420 plates.