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Digestion Problem with BsmBI - (May/05/2006 )

This is my first post (probably of many), so; Hi

I am having trouble digesting my plasmid with BsmBI, I have never used this enzyme before and neither has anyone else I know.

OK, here's the problem, when I digest with BsmBI alone it works but when I do a double digest with XhoI something goes wrong and the gel shows a strange patchy sort of band. The XhoI cuts fine too, when alone.

I was wondering if anyone else has had trouble with this enzyme, and can give me any tips.

Thanks, Erin

-pyro furio-

What temp. are you digesting BsmBI? I think its 55C.

Also you cannot digest with BsmBI for a long time. try to limit the amount of time you are digesting with this enzyme.


ok few questions to start :
did you try a sequential digestion?
do you digest with both enzymes? so in that case do you do a *2step digestion or 1time digest? I mean by a 2 step few h at 37 and then get to 55° (you can use a pcr apparatus for doing that overnight)?
what buffer do you use?


Thanks for your replies, we gave up on BsmBI and are now using another enzyme which is working fine smile.gif


-pyro furio-