If pBR322 cloning vector is cut by Pvu I enzyme for the formation of rDNA then which of its marker gene helps in the selection of recombinants ?

Options

A. `amp^(R) ` gene
B. `tet^R` gene
C. gal gene
D. Both (A) and (B)

Answer

To solve the question regarding which marker gene in the pBR322 cloning vector helps in the selection of recombinants when cut by the Pvu I enzyme, we can follow these steps:

### Step-by-Step Solution:

1. **Understanding pBR322 Vector**:
- pBR322 is a plasmid vector that contains two selectable marker genes: one for ampicillin resistance (bla) and one for tetracycline resistance (tet).

**Hint**: Remember that selectable markers are used to identify which cells have taken up the plasmid.

2. **Identifying the Restriction Site**:
- The question specifies that the vector is cut by the Pvu I enzyme. This enzyme cuts at a specific site within the ampicillin resistance gene.

**Hint**: Look for the specific site where the Pvu I enzyme acts, as this will be crucial for understanding the outcome of the insertion.

3. **Insertional Inactivation**:
- When the gene of interest is inserted into the ampicillin resistance gene (due to the cut made by Pvu I), it leads to insertional inactivation of that gene. This means that the transformed bacteria will lose the ability to grow in the presence of ampicillin.

**Hint**: Insertional inactivation is a key concept in recombinant DNA technology; it indicates that the gene has been successfully inserted.

4. **Survival in Selective Media**:
- After transformation, the bacteria are plated on media containing ampicillin and tetracycline.
- The bacteria with the recombinant plasmid (which has lost ampicillin resistance) will not survive on ampicillin media.
- However, they will survive on tetracycline media because their tetracycline resistance remains intact.

**Hint**: Consider how the survival of bacteria in different media helps in identifying recombinants.

5. **Selection of Recombinants**:
- By comparing the growth on both media:
- Colonies that grow on tetracycline media but do not grow on ampicillin media are identified as recombinants.
- Colonies that grow on both media are non-recombinants (as they have not undergone insertional inactivation).

**Hint**: The ability to differentiate between the two types of colonies is crucial for selecting recombinants.

6. **Conclusion**:
- Therefore, both marker genes (ampicillin and tetracycline resistance) are involved in the selection process, but the tetracycline resistance gene is particularly useful for identifying recombinants after the insertion event.

**Final Answer**: Both A (Ampicillin resistance) and B (Tetracycline resistance) can be used for selection, but specifically, the tetracycline resistance helps in identifying the recombinants.