PCR cloning...help!PCR cloning
Posted 28 March 2012 - 11:05 AM
when I amplify the gene with a cheap taq (non high fidelity) used for genotyping I get my band, no problem.
Now when I try to amplify with high fidelity taqs (Pfu, platinum taq and Phusion polymerase)... non of them work!!!
I have never seen something like this before and its driving me crazy. I also tried to vary some PCR conditions. for Phusion polymerase for instance I've tried with and without DMSO (was thinking maybe the template has some weird 2ndary structures) and I've tried different Mg concentrations for the platinum taq. no luck no band...
any help is appreciated!
Posted 28 March 2012 - 12:19 PM
Posted 28 March 2012 - 05:41 PM
Posted 28 March 2012 - 06:09 PM