help with 5' race
Peter J. Neame
pneame at com1.med.usf.edu
Sun Sep 25 13:33:22 EST 1994
Eric C. Anderson (anderson at pharmdec.wustl.edu) wrote:
: i have tried both the Gibco kit and one from CLONTECH. both of these kits
Ditto on the Clontech kit. We've used it for a couple of proteins where
we wanted to check that the N-terminal wasn't preceded by anything other
than a signal sequence. In both cases, we got about 200 bp of sequence
upstream of the N-terminal. Our 3' primers were in the region 100 -200 3'
of the N-terminal and were not significantly degenerate.
One caveat; the 3' end of the PCR product is liable to be the best
quality. We have only sequenced 3' -> 5' (direct sequencing of the PCR
product) due to "fuzziness" for the PCR band. Of course, if you clone the
PCR product ....
USF and SHCC
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