RT for PCR

[Dr Michael J Carter]

,
With reference to whether RT is the limiting step
in a PCR for long products:

Well we find that it often is. If you know the sequence
a good way to check is to use the 3' primer to make a cDNA
and then PCR the product with the 5'end primer, and a second
about 200 bases from the desired 5'end.

If you get the 200bp product, the RT 3' end was almost
certainly connected and the problem is probably PCR.

Cheers

Mike Carter