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I am trying to optimize annealing temperature for some primer pairs. I have tried optimization using cDNA, genomic DNA, Taq polymerase, phusion polymerase etc., but every time I am either getting non-specific amplification, a band of larger size than expected or no amplification. Every time I am getting a primer-dimer. Can anybody help me optimize the annealing temperature?

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Posting the primer sequences and what genome they are designed against would be helpful. Furthermore, have you double checked that your primers are in the correct orientation? – GWW Aug 15 '12 at 18:29
I've heard of people using single stand binding protein to reduce primer dimers. – Thomas Ingalls Aug 16 '12 at 1:03

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