max. efficient ligation

Netscape User Mika.K.Rantanen at Helsinki.Fi
Tue Apr 2 05:11:39 EST 2002


Hi,
I try to get a maximal ligation efficiency. I have a 900 Bp fragment,
BglII-SalI that is definitely cut at proper sites. My vector has BamHI
and XhoI (definitely cut) and I try to get as many colonies as possible.
I have performed ligations at 16 degrees for 3 hours. Does anyone know
how to increase the ligation efficiency? Is a say 90 % efficiency
obatainable? Can I test this on gel? At the moment my best result is 8
thousand colonies with 25 ng vector, approx equal V:I ratio, TE=10 exp 8
but I want more.
Thanks, Mika


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