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problem with my PCR - primer dimer. - (Apr/18/2005 )

I'm trying to do 3'RACE. (I have already got the 5' end using specific primers). following the RT-PCR using an olido dT, I did PCR with my Specific primer (17bp) and anchor. only the PCR reaction using the anchor primers alone showed a product, however using both primers did not show the product I need except a strong 80bp band (primer dimer??). My product size should be around 200-300bp.
I thought that my DNA copies are very low, or the primers form a dimer.
I have already tried: hot start PCR, touchdoun PCR, gradient PCR and PCR using 50 cycles, and specific different primers.
what else can I try???
thanx for any help!!!!!!!
gall

-gall-

It seems that your GSP didn't work well in your 3'RACE. Why not design another GSP and use the current GSP as the nested primer? so if you have a new GSP ( The expected 3'RACE Product should be longer than 300 bp---which is the expected size of your current GSP with 3' Universal primer), you could use the PCR product as the template and amplify again with the nested primer ( to be exactly, the current GSP and 3'UP). It's normal that you got a few unspecific product from RACE. so it's important to use the nested primer to increase the specificity. good luck.

-littlecell-