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blunt end restriction out of recognition site

blunt end restriction out of recognition site - Molecular Cloning Forum

blunt end restriction out of recognition site - DNA cloning forum. Discuss the cloning of plasmids, vectors, DNA, RNA, cDNA, proteins, and other molecules for expression, sub-cloning, and other methods. Also discuss digestion, ligation, transformation, plasmid preparation, cDNA library screening, and competent cells here.


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Old 05-08-2012, 06:26 PM
Pipette Filler
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Default blunt end restriction out of recognition site



Hi all,

I'm looking for a restriction enzyme that will make blunt cleavage out of it's recognition site.

For example:
if the recognition site is: CAAT
and the sequence is: CAGACAATGCT
GTCTGTTACGA
The resulting cleavage will be something like: CAGA....CAAT....GCT
GTCT....GTTA....CGA

Can any one help me out?
Thank!
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Old 05-09-2012, 03:18 PM
Pipette Filler
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Default Re: blunt end restriction out of recognition site

I don't think something like that exists. Why can't you use normal blunt cleavage? Alternatively, if you are talking about digesting a plasmid, you can run a long PCR with primers terminating with (in your example) 5'-GCT.....-3' and 5'-TCTG....-3'. This way you will get the desired blunt linear DNA (provided you use proofreading polymerases) missing the CAAT fragment.
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