Q.44 Given below are two statements: one is labelled as Assertion-A and the other is labelled as Reason-R

Assertion-A: Observation of cyclosis or cytoplasmic streaming is an indication of active metabolism for checking viability of the freshly isolated protoplasts.

Reason-R: Inclusion of Evan’s blue dye by intact membranes is also indication of viability of the freshly isolated protoplasts.

In the light of the above statements, choose the most appropriate answer from the options given below:

1. Both A and R are correct and R is the correct explanation of A
2. Both A and R are correct and R is NOT the correct explanation of A
3. A is correct but R is not correct
4. A is not correct but R is correct

   Both A and R are correct but R is NOT the correct explanation of A. Cyclosis indicates active metabolism in viable protoplasts, while Evan’s blue exclusion by intact plasma membranes signals viability, but these represent independent tests.

   Question Analysis

   Protoplast viability assessment post-isolation uses multiple criteria. Assertion A correctly links cytoplasmic streaming (cyclosis) to metabolic activity requiring ATP and intact cytoskeleton. Reason R reverses standard dye exclusion logic—Evan’s blue enters damaged cells, so exclusion (not inclusion) indicates viability.

   Option Breakdown

   • Both correct, R explains A: Incorrect. R describes a separate membrane integrity test unrelated to cyclosis mechanism.

   • Both correct, R NOT explanation: Correct. Both valid viability indicators via distinct physiological criteria—metabolism vs. membrane function.​

   • A correct, R incorrect: Incorrect. Despite phrasing ambiguity, standard interpretation confirms R’s viability principle (dye exclusion).​

   • A incorrect, R correct: Incorrect. Cyclosis observation definitively proves active protoplast metabolism and viability.​

   Correct Answer: Both A and R are correct and R is NOT the correct explanation of A.

   Observation of cyclosis or cytoplasmic streaming protoplast viability confirms active ATP-dependent metabolism post-isolation. This dynamic movement along actin-myosin tracks distinguishes living from dead cells.​

   Viability Tests Compared

   Cyclosis: Microscopic visualization of cytoplasmic circulation requires intact cytoskeleton, plasma membrane, and metabolic competence—gold standard for functional viability.

   Evan’s Blue: Vital dye excluded by intact membranes (viable) but taken up by compromised ones (non-viable), staining dead cells blue. Standard exclusion assay.​

   Test Method	Viability Indicator	Mechanism	Equipment
   Cyclosis	Streaming observed	Active metabolism ​	Light microscope
   Evan’s Blue	Dye exclusion	Membrane integrity ​	Light microscope
   FDA	Green fluorescence	Esterase activity	Fluorescence scope
   PI	Red nuclear stain	Membrane damage	Fluorescence scope

   Protocol Application

   Fresh protoplasts (>70% viability needed) show rapid cyclosis within hours post-isolation. Evan’s blue added at 0.25% w/v; viable cells remain unstained after 5-10 min. Both essential for fusion/ transformation success in plant biotech. Critical for CSIR-NET biotech sections.