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Question #47738
The Question
During a recombinant DNA experiment, a researcher fails to elute a specific gene fragment from an agarose gel because the restriction endonuclease failed to cleave the isolated plasmid at its recognition sequence, despite optimal buffer and temperature conditions. Which of the following correctly identifies the molecular cause of this in-vitro cleavage failure and its closest in-vivo parallel in Agrobacterium tumefaciens-mediated plant transformation?
Options
- A.
The recognition site was blocked by host-mediated DNA methylation; analogous to a mutation in the T-DNA border sequences preventing recognition and nicking by Vir endonucleases.
- B.
The plasmid DNA was highly supercoiled, preventing enzyme binding; analogous to the absence of the 'ori' site in the T-DNA preventing its independent replication inside the plant nucleus.
- C.
The restriction enzyme lacked its required RNA primer for cleavage; analogous to the failure of Agrobacterium to synthesize the virA gene product for T-DNA integration.
- D.
The agarose gel concentration was too high for elution; analogous to the plant cell wall physically blocking the entry of the entire Ti-plasmid.
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Correct Answer
Step-by-Step Solution
Explanation
How the Examiner Thinks
Regulatory_Failure
Understand how NOT, EXCEPT, and logical constraints are used to manipulate exam results.
Concepts Behind This Question
Ethidium Bromide Visualization|Gel Electrophoresis Migration|Non-Coding Strand Utility
Why This Question Is Tricky
Common Mistakes
Conceptual Pivot: Students often memorize that restriction enzymes cut at palindromic sequences but overlook the NCERT-derived concept of the restriction-modification system (methylation protecting DNA).
The question demands synthesizing this in-vitro limitation with the in-vivo mechanism of Agrobacterium, recognizing that Vir proteins act as biological restriction endonucleases that require intact border sequences to excise the T-DNA.
Quick Facts About This Question
Question Information
| Subject | Biology |
| Chapter | Biotechnology : Principles and Processes - Class 12 |
| Concept | Ethidium Bromide Visualization |
| Difficulty | 8 |
| Cognitive Load | 5 |
| Target Solve Time | 120 sec |
| Access | Free |