Stupid Ligation - (Apr/24/2003 )
I have been trying to ligate 400 bp fragment into 10.5 kb vector digested with SalI and dephosphorylated with SAP. But I don't get any colonies. I don't know if Sal I is giving some sort of unexplainable star activity. I have cloned this fragment before in pBluescript after PCR and Sal I digestion. Self ligation before dephosphorylation did work once. Can any one give me suggestions? Thanks.
a 11 kb plasmid is not easy to transform, if u didn't get any clone, it may be the reason of low transformation efficiency.u'd better prepare a new electroporated competent cell.