T4 RNA Ligase

Ulf S. Andersson u_andersson2 at yahoo.com
Wed Mar 1 12:12:02 EST 2000


Don't worry about hexamine cobalt chloride (I'm assuming that you are doing
some sort of 5'-RACE). I did the type of ligation you are talking about
recently using only the 10xbuffer provided with the buffer (50 mM Tris-HCl
(pH 7.8), 10 mM MgCl2, 10 mM DTT and 1 mM ATP). I precipitated the oligo and
ssDNA together and performed the ligation reaction in a minimal volume over
night at 37C using 40 units of RNA ligase. It worked fine. But in other
application I don't know whether hexamine cobalt chloride makes any
difference...


Marieluise Kirchner wrote:

> used anyone the T4 RNA ligase for ssDNA / oligonucleotide-ligation? in
> the buffer of my protocoll hexamine cobalt chloride is used. Why? Is the
> effect really that big? Has hexamine cobalt chloride any bad effects in
> further reactions like PCR or ligation into a vector?
> The question is - should i use this hexamine cobalt chloride or better
> not?
>
> thanks and a nice evening
>
> marie
>
> ---





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