I got pCHF3 vector from someone but it is digested with BamH1/Sal1. What I want to do now is to self-ligate the vector to clone a gene using different enzyme sites. What should I do, I know! but I need some suggestion because I might be wrong.
1-Let the vector self-ligate in the presence of T4-DNA ligase or some other ligation solution, how much should be the reaction volume? say for example
Vector 2.0uL+3uL Ligation solution?
2-What verification step I can do after E-coli transformation to confirm that the vector is successfully self-ligated? Maybe I should run both digested and self-ligated vectors to see the difference?
Any suggestion will be highly appreciated ...
Edited by Signal, 20 June 2009 - 04:10 AM.