Practice question

Electrofusion induces protoplast fusion by applying controlled electrical pulses that create reversible membrane breakdown at points of contact allowing lipid bilayers to merge and cytoplasms to coalesce, offering higher fusion frequency viability and selectivity than chemical PEG method. Protocol consists of two electrical phases: first low strength high frequency alternating current 50 to 200 kilohertz field causes dielectrophoresis polarizing protoplasts forming induced dipoles aligning them in pearl chains bringing membranes into intimate contact at poles. Second short high intensity direct current pulses 0.5 to 2 kilovolts per centimeter lasting microseconds generate transient pores dielectric breakdown in contacting membranes that upon resealing fuse into single membrane encircling two protoplasts. Post fusion alternating current field maintained briefly stabilizes heterokaryons. Advantages include ability to monitor individual pair fusion under microscope enabling one to one fusion selection, reduced chemical toxicity preserving division capacity, and high heterokaryon yield up to 60 percent. Parameters voltage duration number of pulses optimized per species to avoid Joule heating irreversible electroporation and cell death. Electrofusion combined with fluorescence activated sorting and microfluidic devices now facilitates precise cybrid and somatic hybrid production for citrus potato Brassica rootstock improvement and organelle transfer programs.