NEETZoologyBiotechnology - Principles and Processes
A student sets up a Polymerase Chain Reaction (PCR) mixture containing template DNA, primers, dNTPs, and Taq polymerase. However, the student incorrectly programs the thermal cycler to alternate only between 54^ C and 72^ C , completely omitting the 94^ C step. What will be the most immediate reason for the failure of DNA amplification in this setup?
Options
- ATaq polymerase will denature at 72^ C and lose its catalytic activity.
- BThe primers will bind to the double-stranded DNA but extension will not occur.
- CThe new DNA strands will be synthesized but will immediately degrade.
- DThe primers will fail to anneal because the template DNA remains double-stranded.
Correct answer
D. The primers will fail to anneal because the template DNA remains double-stranded.
Step-by-step solution
In a standard PCR cycle, the first step is denaturation, which requires heating the mixture to around 94^ C to break the hydrogen bonds between the two strands of the template DNA. If this high-temperature step is omitted and the cycle only alternates between 54^ C and 72^ C , the template DNA will remain double-stranded. Primers require single-stranded DNA to recognize and bind to their complementary sequences. Because the DNA remains double-stranded, the primers will fail to anneal during the 54^ C step. Without