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After the initial primers you selected did not result in the desired product, you spend considerable effort designing two new primers. The only way to avoid he formation of stable primer dimers is by selecting two primers with different annealing temperatures. The forward primer has a Tm of 67C and the reverse primer a Tm of 57C. In order to have a reasonable chance of obtaining the intended PCR product, which temperature should you use for the annealing step in your first PCR attempt with these primers?
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