cloning 2 PCR products into pGEM-T easy vector - (Sep/13/2011 )
I have been trying to ligate 2 PCR products into the pGEM-T easy vector.
PCR product 1 is 13.6 kb long and is digested with BamHI on the 3' end.
PCR product 2 is 2.45 kb long and is digested at the 5' end.
The order in which I want to ligate them is
pGEM-T vector + PCR pdt 1 + PCR pdt 2
So far I have tried 1:1:1 and 1:2:2 ratio for ligation.
None of them have worked for me.
Has anyone else tried this kind of ligation before?
Any suggestions will be greatly appreciated.
I would recommend ligating the two products together first, then try to clone them into the vector.
Also, your products are huge - you may need a bigger vector type such as a YAC or BAC.