Restriction digest - volume of restriction digest (Oct/27/2006 )
I was wondering if anyone knows if the volume in which you do your restriction digest matters. I usually go for 30-50ul reaction with 1ug of DNA but this time I want to digest, heat inactivate, SAP and use the vector including the cut out piece for ligation. I also want to do electroporation and can't have high volume=salt concentration in my ligation. Therefore I wanna digest in 8ul. Any suggestions?
Thanks
hummel
-hummel-
QUOTE (hummel @ Oct 27 2006, 09:48 PM)
I was wondering if anyone knows if the volume in which you do your restriction digest matters. I usually go for 30-50ul reaction with 1ug of DNA but this time I want to digest, heat inactivate, SAP and use the vector including the cut out piece for ligation. I also want to do electroporation and can't have high volume=salt concentration in my ligation. Therefore I wanna digest in 8ul. Any suggestions?Thanks
hummel
What matters is the amount of glycerol in your digest reaction that shouldn't be more than 10%
-dnafactory-
as u wish , but as dnafactory suggested, take care of the glycerol content.
I usually use 20ul or 50ul (for more DNA as I would use more enzyme).
-scolix-