While doing the PCR amplification of metagenomic DNA, I got stuck with no visible bands on gel. But there is always the presence of primer dimers in the gel. I had added about 2.5ul of DNA templte to 25ul reaction mixture, I had added DMSO, Mg2+ etc even all reagents needed for. What could be the possible reason of no band formation? Is it due to less amount of template DNA? or due to any of the PCR components? Does Primer Dimer formation is due to defective PCR reagents? please suggest anything
hi Ahmad Ali ,
primer dimerization is due to internal and dual hybridization of the primers. as in this picture taken from Oligo7, dimerization is avoided by use of DMSO as you did or higher (1-2°c) TM.
fred
Dear Ahmed,
The primer dimer is caused by the primers themselves. You need always to double check when designing your primers that there is no primer dimer possibility. However, using DMSO could help to minimize the dimerization but it still depends on the primers mainly!
Best wishes,
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