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Question #45234
The Question
A researcher attempts to amplify a target gene from an extreme thermophilic bacterium (genomic GC content = 78%) using standard PCR primers that have a GC content of 35%. Despite using Taq polymerase and standard PCR cycles, the reaction yields multiple non-specific bands. Which of the following thermodynamic principles best explains this molecular failure?
Options
- A.
The low GC content of the primers results in a low melting temperature (Tm), causing non-specific annealing to mismatched sequences at standard annealing temperatures.
- B.
The high GC content of the template DNA requires a lower denaturation temperature, leading to template degradation at the standard 94°C.
- C.
The low GC content of the primers increases their affinity to form highly stable primer-dimers via triple hydrogen bonds, depleting the reaction.
- D.
Taq polymerase lacks the helicase activity required to break the dense triple hydrogen bonds of the high-GC template during the extension phase.
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Correct Answer
Step-by-Step Solution
Explanation
How the Examiner Thinks
Comparative_Analysis
Sharpen your ability to evaluate multiple scenarios and distinguish between competing potential answers.
Concepts Behind This Question
DNA|RNA|Tm (Melting Temperature)|Thermophilic Microbes
Why This Question Is Tricky
Common Mistakes
Conceptual Pivot: The direct correlation between GC content, hydrogen bonding, and melting temperature (Tm) in PCR primer design.
The trap relies on two common misconceptions: first, confusing in vivo DNA replication (which uses helicase) with in vitro PCR (which uses thermal denaturation); second, failing to recognize that low GC primers have low thermodynamic stability, leading to promiscuous (non-specific) binding rather than failing to bind at all.
Quick Facts About This Question
Question Information
| Subject | Biology |
| Chapter | Biomolecules - Class 11 |
| Concept | Nucleic Acid Structural Stability |
| Difficulty | 8 |
| Cognitive Load | 5 |
| Target Solve Time | 120 sec |
| Access | Free |