UOG00113@VM.UOGUELPH.CA (craig) (07/28/90)
Hi I'm doing a PCR reaction with an 18 and a 17mer which is giving me problems- primer dimer! I've tried the standard remedies such as changing the annealing temperatures, magnesium concentration, oligomer concentration, template concent ration, etc. I've even used Stratagene's PCR enhancer (doesn't work). I'm usin g BRL Taq polymerase, 25 cycles of 94 C, 55 C, 72 C, with some slight modificat ions, 10-100ng polyA RNA, 10-200ng oligomers, 0.5-2.5mM MgCl2, standard PCR con ditions. The funny thing is that when I use a purified cDNA insert (i.e. omit the reverse transcription step with polyA RNA)I have solved the problem by alte ring my oligo concentrations. No such luck when using message. Any and all sugg estions are welcome. Craig Berezowsky, Dept.Molecular Biology, University of Guelph,Ontario,Canada uog00113@VM.UoGuelph.CA