Nurturing Life Sciences
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F factor
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#Unit 13. Methods in Biology
The two primers used in a PCR reaction have identical Tm for successful and specific amplification.
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The efficiency of amplification declines with an increase in the length of the target sequence.
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Efficiency of PCR is affected by primer length; it declines if the primers used for PCR arc too long.
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The annealing temperature has a marked influence on PCR efficiency; a temperature higher than the ideal annealing temperature reduces PCR efficiency.
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Primer sequence also· affects PCR efficiency. For example, if the primers used for PCR have complementary regions, primer dimers will be formed, and PCR efficiency will markedly decline.
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The sequence of the target DNA itself can affect PCR efficiency. The GC-rich sequences in target DNA may form secondary structures in the single strands produced by denaturation; this could reduce PCR efficiency.
