Help for purification of very short dsDNA

Discussion of all aspects of biological molecules, biochemical processes and laboratory procedures in the field.

Moderators: honeev, Leonid, amiradm, BioTeam

Post Reply
steini2
Garter
Garter
Posts: 1
Joined: Thu Oct 26, 2006 11:03 am

Help for purification of very short dsDNA

Post by steini2 » Thu Oct 26, 2006 11:08 am

Hi everyone,

i could really need some help as i have to separate labeled dsDNA from ssDNA. The fragments are only 40bp long and everything has to be non-denaturating, so it's a bit tricky...
My approach is to try to digest the ssDNA with nuclease S1 and then separate the mononucleotides from the dsDNA by size exclusion chromatography. Any better ideas?

Well, the first experiment didn't work. I still have to work out why. So, here some more questions:
What conditions do you usually use for S1 digestion?
Does pH 4.5 denaturate very short DNA strands?
What if my centrifuge does not go as slow as the required 1000xg for the chromatography spin columns?

I'd be thankful for any help!!

Chris

Post Reply

Who is online

Users browsing this forum: No registered users and 2 guests