Directional cloning

newsnet customer customer at newsnet.com
Mon Mar 27 03:30:53 EST 2006


Hi,

I performed directional cloning of a gene into a vector. 

I was wondering if you need to de-phoshorylate the ends even with directional cloning where the cut vector does not have compatible ends?

It would seem possible that a cut vector could ligate with another cut vector (not self ligate), so it would be a good idea to de-phoshorylate the ends when doing directional cloning?

What are peoples opinion? I just screen about 10 colonies and they are all negative, so there must be some form of re-ligation happening.

Cheers,
Sharp


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