采用基因工程方法对酿酒酵母进行代谢改造，使酵母产生乳酸代谢途径。将来源于 L． mesenteroides和E�coli的D 乳酸脱氢酶基因，分别插入带有G418抗性的酵母穿梭质粒pYX212 kanMX上，电转化酵母，得到2株生产D 乳酸的酿酒酵母重组菌S� cerevisiae WE1510和S� cerevisiae WB1186。进一步摇瓶发酵试验表明：重组菌S� cerevisiae WB1186在YEPD培养基、20 g／L糖、pH 5的条件下生长条件最好，并具有更好的产乳酸能力。经3 L发酵罐条件下验证，S． cerevisiae WB1186分批发酵96 h，最终乳酸积累量达到18�0 g／L；发酵条件为培养基YEPD，接种量10％，溶解氧（DO）30％，转速150 r／min，初始葡萄糖质量浓度10 g／L，控制pH 5�0，通气量3 L／min，OD600最大值转为厌氧发酵。%A Saccharomyces cerevise strain was constructed by genetic engineering method,with lactic acid metabolic pathways. The gene dldh,encoding a lactate dehydrogenase from L. mesenteroides and E. coli, was cloned into a yeast shuttle vector PYX212⁃kanMX. The resultant plasmid pYX212⁃kanMX⁃DLDH was introduced into W303⁃1A by electroporation method and obtained two recombinant S. cerevisiae WE1510 and S. cerevisiae WB1186. Subsequently, a recombinant D⁃lactic acid producing yeast WB1186 was obtained,which had a better ability to produce D⁃lactic acid. It was up to 18�0 g/L of the lactic acid accumulation by batch fermentation in a 3⁃L fermenter for 96 h and the preliminary fermentation conditions were as follows, culture medium YEPD, inoculating rate 10%, DO 30%, stirring rate 150 r/min,concentration of glucose 10 g/L,pH 5�0, aeration rate 3 L/min,when OD600 up to maximum, the fermentation was turned to anaerobic.
展开▼
