plasmid repair

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

On Fri, 2 Jun 2000 14:29:38, "Dr. Duncan Clark" <Duncan at>

= >If both strands are methylated or
ð >unmethylated, the distribution of reversion is 50:50.
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.

| Dr. Peter Gegenheimer       | Vox: 785-864-3939  FAX: 785-864-5321   |
| Department of               |   PGegen at              |
|   Molecular Biosciences     |       |
| University of Kansas        |"When you have excluded the impossible, |
| 2045 Haworth Hall           |  whatever remains, however improbable, |
| Lawrence  KS  66045-2106    |  must be the truth."      S. Holmes    |

More information about the Methods mailing list