User:Jose Fabricio Lopez/Notebook/Logbook/2011/09/04

iGEM 2011

Main project page Previous entry      Next entry

Goal PCR for trouble shooting about PCRs for isothermal assembly. Troubleshooting PCR with Taq Platinum invitrogen and 57°C for annealing. The objectives of those changes are: Change times for elongation. Change Temperature for anneling. Excellent yield by PCR. Gel Completely unsuccessful We are trying to know what is happening with PCRs. PCR with different parameters So far, we have amplified HydG and PFOR1, since now we are trying a new approach. For that we made a simply PCR, this is without GC buffer and DMSO. PCR is for HydEF1 and HydEF2 ( new diluted plasmid with 10ng/μL ) 55°C 60°C HydEF1 HydEF2 HydEF1 HydEF2 Reagents 55°C 60°C Buffer HF Phusion 10μL 10μL dNTPs (10mM each) 4μL 4μL DNA diluited 10ng/μL 1μL 1μL Phusion .5μL .5μL OligoFwd 5μL 5μL OligoRev 5μL 5μL H2O 24.5μL 24.5μL Gel