From 2012.igem.org
Team:Technion/10_September_2012
Ilya
- Did colony PCR for the pCP+RS+mCherry and pCP+RBS+mCherry clones that Rachel prepared for me. There were some positive results but then I realized that they are useless. The previous insert in the plasmid was also mCherry, so I had no way of telling the difference between the background clones and my clones judging by colony PCR.
- Put starters of the colonies I picked for the colony PCR in order to do a test with BamHI digestion tomorrow to find the correct clones.
- Planned some aspects of the gibson attempts that we are going to do this week with Roee and Hila. Tomorrow we will try the assembly of fragments 4 and 5 with 100bp and 400 bp overlaps with different incubation times in 37 degrees followed by the incubation in 50 degrees which is instructed in the gibson protocol.
- Prepared a 0.6% gel for testing the gel electrophoresis conditions for the gibson results.
Inbal
Asaf
Hila
Lior
Noa
Evgeni
- Due to continuous failure of Bba_015 insertion in pPROLar plasmid I'll try new strategy: instead of double digestion of pPROLar I'll do sequential digestion: HindIII-HF restriction -> Heat inactivation of HindIII-HF at 80C -> BamHI-HF restriction.
- In addition I'll do new Bba_015 restriction with BamHI+HindIII, maybe ligation problems were due to problem with biobrick restriction (Done by Rachel)
- Overnight ligation of newly prepared pPROLar and Bba_015
Shahar
Rachel
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