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This research was focused on investigation of the condition for protoplast formation and regeneration of protoplast fusion between Saccharomyces cerevisiae which has fermentation ability and Candida cariosilignicola which can grow at high temperature and utilize methanol.
The results obtained were as follows;
The highest production was collected in exponential growth phase. Ninety-nine°/. protoplast formation of C. cariosilignicola was obtained in glycin-NaOH buffer (pH10.0) containing Zymolyase 0.5mg/m/ at 35°C for 1hr incubation.
The highest regeneration was produced when protoplast suspension containing 0.5% soft agar in buffered 50mM CaC12 was poured as a soft overlay onto 2% agar plates. Equal amount of protoplast suspension of two strains was mixed and centrifuged. The subsequent pellet was added to 2m1 of 35°7% polyethylene glycol (MW 4,000) containing 50mM CaC12, and incubated at 30°C for 10mia. Then 0.1m1 of the suspension of aggregated protoplast was immediately covered with minimal medium and incubated at 40°C for 5-7 days. As results, SCI, SC2, and SC3 fusants were obtained. The physiological characteristics of fasants produced by protoplast fusion were; SCI, and SC2 utilized maltose, galactose, methanol, potassium nitrate. SC3 utilized all the above materials except galactose.
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