Question

Arrange the steps in PEG induced protoplast fusion in correct sequence -
A. Treatment of protoplast mixture with 28-50% PEG for 15-30 minutes.
B. Protoplast aggregation.
C. Washing of protoplast (alkaline medium pH 9-10), and high Ca\textsuperscript{2+ concentration}
D. Selection of protoplasts of different strains / species.
Choose the correct answer from the options given below :

• A. A, B, C, D
• B. D, A, B, C
• C. C, D, B, A
• D. D, B, C, A

Hint:

Think of the fusion process logically: {Select} your ingredients (D) \(\rightarrow\) {Add} the "glue" (PEG) (A) \(\rightarrow\) Observe them {sticking} together (B) \(\rightarrow\) {Wash} and finalize the fusion (C).

Answer 1

The Correct Option is B

Step 1: Understanding the Concept:
The question asks for the correct chronological order of steps for fusing plant protoplasts using Polyethylene Glycol (PEG) as a chemical fusogen.
Step 2: Detailed Explanation:
The logical protocol for PEG-induced fusion is as follows:
D. Selection of protoplasts of different strains / species. The very first step is to prepare and select the parent protoplasts that are to be fused.
A. Treatment of protoplast mixture with 28-50% PEG for 15-30 minutes. The selected protoplasts are mixed, and the fusogen, PEG, is added. PEG is a dehydrating agent that helps bring the protoplast membranes into very close contact.
B. Protoplast aggregation. As a direct and immediate result of the PEG treatment, the protoplasts clump together, or aggregate. This close contact is a prerequisite for fusion.
C. Washing of protoplast (alkaline medium pH 9-10), and high Ca\textsuperscript{2+ concentration.} After the incubation with PEG, the PEG is carefully washed away or diluted with a special elution solution. This solution has a high pH and a high concentration of calcium ions (\(Ca^{2+}\)). These conditions reduce the negative charge on the protoplast surface and help in the coalescence of the membranes of the aggregated protoplasts, leading to fusion.
Therefore, the correct sequence is D \(\rightarrow\) A \(\rightarrow\) B \(\rightarrow\) C.
Step 3: Final Answer:
The correct sequence of steps is D, A, B, C.