sequencing without precipitation

didier d-fesque at nimr.mrc.ac.uk
Thu Apr 24 04:58:55 EST 1997


hey, have a look to Amersham kit, quick denature sequencing kit (USB
kit). this kit is based on a protocol published in NAR a few year ago
(have a look on medline!). basically , using miniprtep DNA denature 8ul
(500ng-3ug) of plasmid DNA with 2 microl 1M NaOH add 1ul (10ng) primer,
incubate 10 min 37 degre, let sit on ice a few minute. add 2 microl of 1
M HCl , add 2 ul plasmid buffer (unknown composition , from the
kit,absent from the original protocol) incubate 10 min 37 degre, let sit
on ice. sequence as with the sequenase version 2.0 kit.

it work and give very reliable result but sometime it give very faint
bands .

didier



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