Long PCR with degenerate primers and inosine

Rohit Mistry r.mistry at udcf.gla.ac.uk
Wed Nov 22 14:56:43 EST 1995


Hello,
I would be very interested in hearing from anyone who has had 
experience using the Gibco/BRL Elongase PCR kit (containing 
pyrococcus thermostable polymerase and Taq). I have just tried it 
out to amplify a ~12 kb product. However, I could not see any bands 
at all, apart from primers. I'm using  30-mer degenerate oligos. 
Also the primers contain inosine. 

The PCR conditions were those recommended by the kit. Annealing 
temperature of 55 C was used, which is known to give a range of 
shorter non-specific products using Taq alone.

Thank-you



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