摘要
研究细胞培养和溶菌酶酶解条件对罗伊氏乳杆菌原生质体形成率的影响以及罗伊氏乳杆菌原生质体在多种再生培养基中的再生条件,结果表明罗伊氏乳杆菌05-1和05-2原生质体形成的最适条件:在MRS培养基中接种量5%(两菌株活菌数分别为4.96×108 cfu/mL和8.04×107 cfu/mL),37℃静置培养至OD600值1.0(两菌株活菌数分别为2.36×1012 cfu/mL和6.56×109 cfu/mL)左右,酶解pH8.0、酶质量浓度20 mg/mL、酶解时间45min。在此最适条件下,两菌株原生质体形成率分别为90.2%和92.8%;在最适再生培养基RM1中,罗伊氏乳杆菌05-1和05-2原生质体的再生率分别为30.6%和23.2%;RM1中,胎牛血清(BSA)是罗伊氏乳杆菌原生质体再生不可获缺的物质。本研究为构建益生乳杆菌食品级基因克隆和表达系统,实现原生质体的转化及细胞融合育种与基因组洗牌分子育种,提供理论依据。
The effect of protoplasts formation ratio in LactobaciUus reuteri have been researched with cell culture and lysozyme conditions in this paper.Regeneration of protoplasts in Lactobacillus reuteri have been explored on multimedia. The results showed that Lactobacillus reuteri 05-1 and 05-2 protoplast formation of the optimum conditions were as follows：5% of incubation in MRS media （The viable cells counts of two strains were respectively 4.96×10^8 cfu/mL and 8.04×10^7 cfu/mL）, strains were cultured at 37 ℃ with an optical density about 1.0 at 600 nm （The viable cells counts of two strains were respectively 2.36×10^12 cfu/mL and 6.56×10^9 cfu/mL）,range of pH 8.0, enzyme concentration of 20 mg/mL, treating time of 45min, in the optimum conditions, the two strains protoplast formation rates were reached to 90.2% and 92.8% respectively. The ratio of regeneration of Lactobacillus reuteri 05-1 and 05-2 were 30.6% and 23.2% in the optimum renewable medium RM1. In the renewable medium RM1, fetal calf serum （BSA） was the indispensable material to the regeneration in Lactobacillus reuteri. This research provide practical basis, in order to construct of the food-grade gene cloning and expression system for Lactobacillus, realize the protoplasts transformation, and screen good stains by fusion of Lactobacillus and genome shuffling.
出处
《中国食品学报》
EI
CAS
CSCD
北大核心
2010年第1期10-18,共9页
Journal of Chinese Institute Of Food Science and Technology
基金
国家高技术研究发展计划(863计划)项目(No.2006AA10Z317)
关键词
罗伊氏乳杆菌
原生质体
制备
再生
Lactobacillus reuteri
protoplasts
formation
regeneration
