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#protoplast isolation

3 public questions tagged with this topic.

Plant tissue culture 2

Osmotic stabilizers such as mannitol are required to:

Osmotic stabilizers such as mannitol sorbitol glucose at 0.3 to 0.7 molar act as essential protectants preventing protoplast bursting after cell wall removal because wall normally provides mechanical resistance counterbalancing internal turgor pressure generated by high intracellular solute concentration. Without rigid wall plasma membrane alone insufficient to resist water influx driven by lower external water potential, protoplasts swell and lyse within minutes, destroying preparation. Non metabolizable sugar alcohol mannitol provides iso osmotic environment matching cell sap osmolarity abou

Ref: Kao & Michayluk 1975 osmotic stabilizer; Plant Cell Culture protoplast methods.

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Plant tissue culture 2

Most common tissue source for protoplast isolation is:

Fully expanded young leaves represent most common and convenient source for protoplast isolation across many species due to favorable physiological and structural attributes maximizing yield and viability. Young leaves have thin primary walls composed of cellulose hemicellulose and pectin not yet extensively lignified or suberized, allowing efficient enzymatic digestion within 4-12 hours using cellulase plus macerozyme mixture. Mesophyll cells are large highly vacuolated with minimal intercellular air spaces ensuring enzyme access, contain chloroplasts providing visual marker for intact protop

Ref: Evans & Bravo 1986 mesophyll protoplast; Bhojwani & Dantu leaf protoplast protocol.

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