Sequencing high G + C DNA

Allen Gathman C741SCB at SEMOVM.SEMO.EDU
Tue May 2 11:34:56 EST 1995


In article <3o5dk4$rv3 at synapse.bms.com> Steven Goldberg <goldberg at bms.com> writes:
>I have been attempting to sequence DNA of high G + C content (ca. 72%)
>.

(snip, snip)

We have similar problems.  Although we haven't yet figured it out, we
are about to try the suggestions in a paper titled "General method for
amplifying regions of very high G+C content" from NAR 21:2953-2954.

Basically, the authors say to use denaturation temp of 98 C, five min
combined annealing/elongation at 70 C, and to try reducing template
concetntration.  (Obviously, I'm assuming you're using a cycling
sequencing protocol here.)

We're about to try this; will let you know.  I'd appreciate hearing any
other suggestions.

************************************************************************
|    Allen Gathman                |                                    |
|    Biology Department           |  Internet: C741SCB at SEMOVM.SEMO.EDU |
|    Southeast Missouri St. U.    |  Phone: (314) 651-2361             |
|    Cape Girardeau, MO 63701     |  Fax: (314) 651-2223               |
************************************************************************
|    "Anyone would know that," said Dorothy.                           |
|    "Certainly; that is why I know it," returned the Scarecrow.  "If  |
|    it required brains to figure it out, I never should have said it."|
|    -L. Frank Baum                                                    |
************************************************************************



More information about the Methods mailing list