Hot Alkaline Minipreps for automated sequencing

Philip L. Carl plc at med.unc.edu
Fri May 13 14:28:54 EST 1994


We have encountered a very strange problem when trying to use the Hot
Alkali Miniprep method of Musich and Chu Biotechniques 14:958 (1993).  The
DNA does not seem to precipitate in the final isopropanol step unless we
put in a chloroform extraction step.  The final DNA we got didn't sequence
at all, but that may be another problem of deletions of the priming sites.
  However, I was wondering if others have tried this technique that is
claimed to be very robust for automated fluorescent sequencing, and if so
how it compares in your hands to procedures such as Qiagen columns and PEG
precipitations.  By the way, the A280/260 of DNA prepared by this method
is very low although the authors say this doesn't hurt sequencing.  After
a single chloroform extraction, it looks much better, but as I said-no
sequencing.

Thanks for any help.

Phil Carl



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