Question

A plasmid vector contains a multiple cloning site (MCS) within the lac-Z gene. If foreign DNA is inserted into the MCS, what happens when competent cells are transformed with this plasmid and allowed to grow on a nutrient medium plate with X-gal and IPTG?

• A. Transformed cells with recombinant plasmids will appear blue.
• B. Transformed cells with recombinant plasmids will appear white.
• C. All transformed cells will appear blue.
• D. Non-transformed cells will appear white.

Hint:

Remember the key idea of blue-white screening:

\textbf{Blue} = Bad (for the experimenter). The plasmid is non-recombinant.
\textbf{White} = Wanted. The plasmid is recombinant, containing your gene of interest.
The insert inactivates the "blue-making" gene.

Answer 1

The Correct Option is B

Step 1: Concept Overview:
Blue-white screening identifies bacteria with recombinant plasmids (plasmids with foreign DNA inserts). This method uses insertional inactivation.
Step 2: Detailed Explanation:
Key components:

lac-Z gene: Encodes \(\beta\)-galactosidase.
X-gal: A colorless \(\beta\)-galactosidase substrate. Cleavage yields a blue product.
IPTG: Induces the lac operon, ensuring lac-Z gene expression.
Multiple Cloning Site (MCS): A region with unique restriction sites within the lac-Z gene.
Transformed cell outcomes:

Non-recombinant plasmid: Plasmid re-ligates without an insert, leaving the lac-Z gene intact. Functional \(\beta\)-galactosidase is produced, cleaving X-gal, resulting in blue colonies.
Recombinant plasmid: Foreign DNA inserts into the MCS, disrupting (insertional inactivation) the lac-Z gene. No functional \(\beta\)-galactosidase is produced. X-gal is not cleaved, and colonies remain white.
Non-transformed cells (no plasmid uptake) won't grow on antibiotic-containing medium if the plasmid carries an antibiotic resistance gene. If they did grow, they would be white due to lacking the lac-Z gene.
Step 3: Conclusion:
Transformed cells with recombinant plasmids, having an inactivated lac-Z gene, appear white on X-gal/IPTG medium.