Re Bcl1

Wed Sep 16 13:44:40 EST 1992

mdv6n at (Mark D. Van Cleve) writes:
>Anyone else have trouble with this enzyme?  Either it goes bad very quickly
>in our freezer, which would be odd for a heat stable enzyme, or we're not
>using it correctly, or something, because BclI from several different 
>sources has very little interest in cutting plasmid for us, even when it
>will consent to cut lambda, which is not always.  Any clues out there?>

	I don't know if this is your problem, but Bcl1 is sensitive to DNA
methylation by the (aptly named) _dam_ methylase. Dam+ _colis_ put a
N6-methyladenine in the sequence GATC, which is part of the Bcl1 recognition
sequence. Won't cut worth a darn in most of the common lab strains. I had this
problem several years ago with some DNA containing synthetic linkers I had made.
The original construct cut fine, but once it was cloned into _coli_ and
reisolated it wouldn't cut at all. Made me feel a little silly, once I figured
out what the problem was.


Mike R.
   | Dr. Michael P. Ready                     | READY at UTBC01.CM.UTEXAS.EDU |
   | Clayton Foundation Biochemical Institute | Phone: (512)471-3625       |
   | University of Texas                      | FAX:   (512)471-8696       |
   | Austin, Texas 78737                      |                            |

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