directional cDNA cloning
myagi at u.washington.edu
Thu Jan 4 14:40:18 EST 1996
I'd like some advice on directional cDNA cloning. In the past I've
successfully constructed nondirectional cDNA libraries, but for a variety
of reasons we're considering doing a directional library in a homemade
vector with EcoRI (5') and XhoI (3') sites. Stratagene sells a kit using
precisely these enzymes, but before popping $600 I'd like to get some
feedback both on the kit and the potential difficulties I might encounter
in the protocol. Essentially, the protocol substitutes methyldC for dC in
the cDNA synthesis reaction, the idea being to prevent cleavage by XhoI
at any internal sites.
I'd appreciate hearing from anyone who's used this kit, and any
suggestions from others who have done directional cloning.
Thanks in advance,
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