ClonTech RACE PCR
Gary E. Vallad
vallad at badlands.nodak.edu
Tue Nov 18 23:03:36 EST 1997
Hi! I'm having some problems with my RACE PCR reactions and hoping that
someone may have some insight. I've been performing 5' and 3' RACE PCR using
cDNA prepared via ClonTech's Marathon cDNA Kit. I am getting some useful
products, but I'm also getting some non-specific products derived from the
linker specific primer binding to complementary regions in my cDNA other than
the cDNA linker. Yet, when I run controls using only my linker specific
primer alone, I get no product. Also, I know my gene-specific primers are
working since when I use both 5' and 3' gene-specific primers in conjunction I
get the appropriate sized fragment. I've been getting these non-specific
products when running a variety of different RACE reactions with different
gene-specific primer sets. I'm assuming that they are just miscellaneous
products isolated along with my fragment of interest in the GeneClean process,
but I've been getting a high percentage of clones that are these non-specific
type of products. Has anyone else had problems with this. Like I mentioned
before my control with just the linker-specific primer comes up with no
products that can be visualized on a gel. Any help would be much appreciated.
Thanks,
Gary E. Vallad
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