Meaning of high MW PCr smears
Steven Sullivan
sullivan at gwis2.circ.gwu.edu
Mon Jun 2 23:03:27 EST 1997
I'm trying to re-amp some PCR products from a gel -- running them in an
EDTA-free gel and buffer, taking a plug, melting it in 20X volume of
distilled water (95 degrees for a few minutes; the gel is *not* low melt),
using 1 ul in a 50 ul PCR reaction, with M13 F and R primers at 55 degree
anneal temp (same as original PCR). ANd I keep getting smears starting
from around the expected product size up to several kb higher. No
primer-dimers in evidence (I'm doing hotstarts). Am I adding too much
DNA, or is there some other parameter that needs changing?
More information about the Methods
mailing list