Sequencing problem. Is it the GeneClean?

Bernard Heymann bheymann at bragg.bio.purdue.edu
Wed Feb 9 10:33:27 EST 1994


In article <94039.133242NORVALC at QUCDN.QueensU.CA>,
<NORVALC at QUCDN.QueensU.CA> wrote:

>   My question is straightforward:  Have anybody else noticed that there are
> critical steps in the GeneClean protocol?  If so, what should I be focusing on?
> 
>   Thanks in advance for your help.  I will summarize and post any replies.
> 
> Barry Campbell

One of the irritating problems with GeneClean I found was that it is
sometimes impossible to avoid carrying over some of the glassmilk after the
final elution. I routinely microfuge the final product again and transfer
the liquid to a new tube. Although I didn't do any controlled tests, I got
the impression that glassmilk in the prep decreases ligation efficiency.
The other problem is that I found recoveries of just about any size
fragment is poor (around 20 %, definitely below 50 %). The other protocol I
have tried is the Magic PCR prep from Promega, with which I doubled my
recoveries and had less trouble with ligation. However, with all the bad
anecdotes flying around about the new Wizard preps, I am getting a little
nervous using a related prep from Promega.

-- 
Bernard Heymann
bheymann at bragg.bio.purdue.edu



More information about the Methods mailing list