BsaBI digests

Bernard Murray bernard at elsie.nci.nih.gov
Tue Jan 16 12:21:41 EST 1996


In article <JDuPvKAm75+wEw4R at genesys.demon.co.uk>, duncan at genesys.demon.co.uk 
says...
>
>In article <v01530501ad207196f61c@[142.20.8.19]>, jlight at RESUNIX.RI.SICK
>KIDS.ON.CA writes
>>Does anyone out there have experience doing BsaBI digests?  It's one of
>>those enzymes with activity that's blocked by overlapping dam methylation.
>>I'm trying to cut a plasmid with a single BsaBI site.  The plasmid was
>>recovered from strain GM2163, which is dam(-) and dcm(-), but the digest
>>doesn't appear to be working any better.  Any advice would be greatly
>>appreciated.
>>
>

Just wondering, did you grow the GM2163 in the presence of chloramphenicol
to maintain the methylation negative status?  I have my stocks selected
at 100 mg/l and plasmids with ClaI in a dam context cut to completetion
when this is used as the host (no cut at all with XL-1 or DH5alpha).

>Can you cut the supposed dam- plasmid with Mbo I. If so it is definitely
>dam-.

This would be a good test of plasmid methylation status and your BsaBI.

	Happy cutting,
		Bernard
Bernard Murray, Ph.D.
bernard at elsie.nci.nih.gov  (National Cancer Institute, NIH, Bethesda MD, USA)




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