Summary: | 碩士 === 國立屏東科技大學 === 食品科學系 === 93 === The purpose of this study tried to breed the starch-fermenting yeast by protoplast fusion. The fusion was carried out between the protoplasts of Schizosaccharomyces pombe and saccharolytic molds such as Asp. niger. The method of protoplasts preparation for removing the cell wall of two parents strains, the cell wall lytic enzyme from Trichoderma viride was used. Then, the protoplasts fusion was promoted by polyethylene glycol (PEG), and cell wall regeneration of fusants was carried out in regeneration medium. So, new yeasts of Fusant were isolated. Thus, the first time isolated fusant was named FusantA1, then, FusantA1 subsequently fused with other saccharolytic mold isolated fusant was named FusantAA2, in the same way, FusantAA2 fused again with other saccharolytic mold isolated fusant was named FusantAAR3. Morphology of three fusants were investigated. The assimilation and fermentation ability of carbon sources, and assimilate ability of nitrogen
were investigated, too. The results showed the amylolytic enzyme activity of FusantA1, FusantAA2 and Fusant AAR3 were 6.2 U/ml, 8.2 U/ml and 10.5 U/ml, respectively. Morphology of these fusants all are sphere; three fusants could assimilate many kinds of carbon source and nitrogen source. In fermentation ability of these three fusants could ferment potato starch and trehalose, but Schiz. pombe without these ability.
The optimum culture conditions of these fusants were the medium contained 2-3%potato starch and 0.5%yeast extract at pH 4 and growing at 30℃.After seed cultured for 24 hr, 48 hr and 72 hr then transferred to a new medium cultured 72 hr, respectively, their amylolytic enzyme activity of FusantAAR3 were 2.9 U/ml, 4.0 U/ml and 12.3 U/ml; after fermented for 20 day, fermentation ability of FusantAAR3 were 58.8%, 59.3%, 75.3%, respectively. Additions of enzyme in ethanol fermentation medium, fermentation ability of FusantAAR3 were accelerated according to the enzyme activity was increased.
The stability of these Fusants were found FusantAAR3 better than FusantAA2 and A1. Enzyme activity of FusantA1 was decreased during storage at 4℃ after 4 months. But, the growth rate and enzyme activity of FusantA1 were promoted by transferring in the medium. When transferring 5 times, the activity of FusantA1 was recovered to their original enzyme activity and growth rate.
|