Detecting Biotinylated PCR products?

Jim Gerlach GERLACHJ at QUCDN.QueensU.CA
Tue Sep 14 10:48:26 EST 1993

In our efforts to do quantitative PCR we have in the past kinased oligos 
with 32P. Recently we tried using oligos with a 5' biotin group. Following 
separation on a gel and alkaline transfer to a nylon membrane we have been 
visualizing bands with a streptavidin-alkaline phosphatase conjugate and  
chemiluminescence (LumiPhos 530). We have also cut out the bands from the 
nylon membrane and measured the light with a luminometer. All of this worked 
reasonably well and we want to continue using the biotinylated oligos.

What I would like to do is eliminate the transfer step. I have considered 
doing the biotin-avidin step directly on gels (dried or semi-dry) or using 
magnetic beads on the opposing oligo to separate PCR product from primers. 
Any experiences or suggestions would be appreciated. Thanx in advance.
Jim Gerlach                               Cancer Research Laboratories
gerlachj at qucdn (BITNET)                   Queen's University
gerlachj at (InterNet)      Room A309, Botterell Hall
tel (613) 545-6446                        Kingston, Ontario
fax (613) 545-6830                        Canada K7L 3N6

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