plasmid repair

Dr. Peter Gegenheimer PGegen at UKans.nolospamare.edu
Fri Jun 2 21:37:07 EST 2000


On Fri, 2 Jun 2000 14:29:38, "Dr. Duncan Clark" <Duncan at nospam.demon.co.uk>
wrote:

= >If both strands are methylated or
ð >unmethylated, the distribution of reversion is 50:50.
ð
ð A further question on this.  Based on the above, because standard (well
ð it used to be years ago!) M13 site directed mutagenesis would have given
ð a mismatched unmethylated strand and presumably the original ssDNA
ð template strand was methylated(*) how come you got mutants? Or is it the
ð case that the mutation frequency was very very low but you still managed
ð to find the mutant by probing.

The efficiency of M13 mutatgenesis by the original methods was never more than
50% (the theoretical limit), and was usually 10-20 percent. As I recall, 30%
was about the upper limit in practice.

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