Why low second-strand cDNA?
graham at biodec.wustl.edu
Wed Apr 24 17:57:18 EST 1996
Comments welcome on the possible causes of low conversion of
good first strand cDNA synthesis to double stranded product.
Of course, the second strand is primed by RNA following RNAse H
cleavage, and may have been degraded during a 1 h first strand
synthesis. Is there a convenient way to rescue this first strand
material (eg. random prime and fill?). Short products are not
of conern, but lack of linker-ligatable product ends certainly is.
Does anyone add additional random primers during the second-strand
synthesis? Is there a problem with this other than reduced
length that I've overlooked? :)
J. Graham PhD
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