Q.
In the polymerase chain reaction, the optimum temperature for the extension step is
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NTA AbhyasNTA Abhyas 2020Biotechnology : Principles and Processes
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Solution:
A single PCR reaction involves three temperature-dependent steps, described as follows: i) Denaturation: The starting solution is heated, usually to 94oC. The high temperatures break the hydrogen bonds between the two strands of the original DNA double helix, providing the necessary single-stranded templates. ii) Annealing: After just a couple of minutes at that temperature, the reaction mixture is quickly cooled, usually to somewhere between 50oC and 60oC. It is then held for less than a minute at this lower temperature which is enough time for the primers to bind to their complementary sequences on the single-stranded templates. iii) Primer extension (polymerization): The sample is next heated to 72oC for some time, during which time the DNA polymerase adds nucleotides to the primer, synthesizing a new DNA strand using only the template sequences that bind the primers.