Nurturing Life Sciences
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#Unit 13. Methods in Biology
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.
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7-dcaza guanosine triphosphate (7-deaza OTP) can be used in place of OTP; it reduces hairpin loop formation without affecting the Watson-Crick base pairing required for annealing and primer extension.
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Addition of certain proteins, such as, bovine serum albumin enhances efficiency by protecting the DNA polymerase and by binding to PCR inhibitors.