problems with restriction digestion

WS via methods%40net.bio.net (by novalidaddress from nurfuerspam.de)
Fri Feb 8 17:00:16 EST 2008


Hi Mary,

can you run your DNA preps on the gel, too?. A too long alkakline step
(some ptotocols say 5min) definitely will kill your plamsid
(partially) and cause it to wind up in some strange undigestable
conformation that may appear as extra band. For standards E.coli lab
strains, a few seconds definitely are enough and you should work on
ice and immediately proceed with neutralization.

Beware, too, of XbaI sensitivity to certain methylation patterns
possibly causing poor or partial digests.

Best regards,

Wo





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