RNAseq circligaseII

[andolSDU]

Hi everbody

I am working with E. coli and I am trying to prepare librabries for Solexa sequencing

I found a protocol for sample prep, were they use CircligaseII to circularize cDNA.
I have tried to circularize cDNA extracted from an acrylamide gel, but it does not seem to work very well.

Does anybody have expience with this enzyme?

When you look at the protocol that comes with the enzyme it looks as though
you do not need to add ATP to the reaction, and it does not seem to be in the reaction buffer, however whith the circligaseI enzyme you do need to add ATP. Do you add ATP to your reactions.

[csc]

One of the main differences between circligase I and II is that circligase II is basically pre-adenylated which allows it to act more efficiently. So, to answer your question, no I don't add ATP to circligase II.

[mheisler]

Do you have a reference for that protocol? I am also interested in using Circligase II for cDNA preparation.

Actually its ok, I've found it....google is my friend.