Is ligation reaction volume only 10 ul?

Frank O. Fackelmayer fof1 at chclu.chemie.uni-konstanz.de
Fri Jun 13 07:26:51 EST 1997


In article <01bc77fe$cd0533c0$467df88f at KAISTWF.kaist.ac.kr>, "Yeon Chul
Kim" <e_yckim at cais.kaist.ac.kr> wrote:

> Hi!!
> 
> In my knowledge, most people used 10 ul of ligation reaction volume.
> 
> Are there some problems using greater than 10 ul (for example 50 or 100
> ul)?
> 
> Please give helpful information.

Hi Yeon,
The reason why most people use small volumes for ligation is that ligation
efficiency depends on the DNA concentration. Thus, there is no problem in
doing ligations in say 100ul, except you wont get as many ligated products
(and clones). You should also keep in mind that self-ligation
(intramolecular ligation) of linear DNA molecules (e.g. your vector) is
preferred over intermolecular ligation (e.g. vector + insert) at low DNA
concentrations.

Hope this helps,
Frank



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