Blunt/Sticky-end ligation

Nicolas Lemee Lemee at
Tue Sep 7 11:50:39 EST 1999

Hello, my two cents :) :
If the enzymes that cut the vector do not produce compatible ends then I
never dephosphorylate the vector.  I always use the plain buffer that came
with the ligase. 3 to 1 insert/vector ratio & room temperature overnight is
what I usually do for all ligations and it works fine with me.

Stefan Kappeler wrote:

> I should ligate a 1kb insert into a dephosphorylated 6.5 kb vector. One
> end is sticky (XbaI) the other blunt (PmlI). What are the best reaction
> conditions:
> -temperature, time
> -buffer composition (ATP, PEG,...)
> -molar ratio insert/vector
> Many thanks
> Stefan

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