Question

Which of the following is not used in protoplast fusion?

• A. $Ca^{++}$ treatment
• B. PEG
• C. High pH
• D. $CsCl_2$

Hint:

The combination of PEG, high $Ca^{2+}$, and high pH is the most efficient chemical method for somatic hybridization, often achieving much higher fusion rates than using any of these agents alone.

Answer 1

The Correct Option is D

Step 1: Understanding the Concept:
Somatic hybridization involves isolating protoplasts by removing the cell wall, followed by fusing protoplasts from different species to generate a hybrid plant.
Because protoplasts carry a net negative surface charge, they naturally repel each other.
Chemical or physical agents, known as fusogens, are required to reduce this electrostatic repulsion and induce membrane fusion.

Step 2: Detailed Explanation:
Let us examine the role of each chemical agent listed in the options.
Polyethylene glycol (PEG) is the most widely used chemical fusogen.
It acts as a molecular bridge, dehydrating the protoplast membrane and bringing adjacent membranes into close contact to facilitate fusion, which matches Option (B).
Treating protoplasts with calcium ions ($Ca^{2+}$) at a high pH (around 10.5) is another standard chemical fusion method.
The high pH alters the membrane charge, and the divalent calcium ions bind to the negatively charged membrane lipids, neutralizing surface charges and facilitating fusion, which matches Options (A) and (C).
Cesium chloride ($CsCl_2$) is a heavy metal salt used primarily in density gradient ultracentrifugation to separate and isolate DNA or RNA fractions based on buoyant density.
It does not possess fusogenic properties and is not used in protoplast fusion protocols.

Step 3: Final Answer:
$CsCl_2$ is not used in protoplast fusion.
This corresponds to Option (D).