关键词: 红发夫酵母; 黏红酵母; 原生质体; 蜗牛酶; 溶菌酶

Abstract: This experiment was conducted to screen astaxanthin-high-producing yeast strains by using protoplast fusion technology to evaluate the optimum parameters of cell age(h), reactive temperature(℃), reactive time(h) and osmotic stabilizers of protoplasts formation and regeneration of Phaffia rhodozyma mutant and Rhodotorula glutinis mutant. The product of protoplast formation rate and regeneration rate were as testing indicators. The results showed that the product of formation rate and regeneration rate in logarithmic growth prophase of Phaffia rhodozyma and that in logarithmic growth metaphase of Rhodotorula glutinis were significantly higher than that in other groups (P<0.05);the product of formation rate and regeneration rate of Phaffia rhodozyma of 22 ℃ was 17.34%,those of 25 and 28 ℃ were 0, those of 22 and 25 ℃ of Rhodotorula glutinis were significantly higher than that of 28 ℃ (P<0.05),but there were no significant differences between these two groups (P>0.05);the product of formation rate and regeneration rate of two strains in the reaction time of 2 hours were significantly higher than those in other groups (P<0.05). The product of formation rate and regeneration rate in the control group were significantly higher than those in other groups (P<0.05).The suitable processing conditions of protoplasts formation and regeneration showed as follows: cell age of Phaffia rhodozyma was 14 h, cell age of Rhodotorula glutinis was 18 h, the reactive temperature was 22 ℃, the reaction time was 2 h, osmotic stabilizer of protoplasts formation was KCl buffer of 1.0 mol/L, osmotic stabilizer of protoplasts regeneration was sucrose diluents of 17%, finally selected one fusion srain that could grow well at 28 degrees and had 586.38 μg/g astaxanthin content, increased by 79.58% and 64.08% compared with their paraent strains, and the fermention tests showed that the ability of producing astaxanthin was still stable after many generations culture.

Key words: Phaffia rhodozyma; Rhodotorula glutinis; protoplast; snailase; lysozyme