摘要
甘油是一种极其理想的耐高渗透压介质。利用PCR方法,从产甘油假丝酵母WL2002-5中扩增出了2个产甘油的关键酶基因GPD和GPP,分别编码3-磷酸甘油脱氢酶(glycerol3-phosphatedehydrogenase,GPD)和3-磷酸甘油磷酸酶(glycerol3-phosphate phosphatase,GPP)。利用T-Vector在Escherichia coliJM109中克隆得到大量的GPD和GPP基因,并成功构建了重组质粒pYX212-GPD和pYX212-GPP;通过LiAc转化法将重组质粒导入酿酒酵母Saccharomyces cerevisiaeW303-1A。初步实验结果表明发酵过程中pYX212-GPD/S.cerevisiaeW303-1A的生物量高于pYX212-GPP/S.cerevisiaeW303-1A和野生型S.cerevisiaeW303-1A;发酵72h后,pYX212-GPD/S.cerevisiaeW303-1A发酵液中甘油含量大约为12mmol/L,明显高于野生型S.cerevisiaeW303-1A的甘油含量,而pYX212-GPP/S.cerevisiaeW303-1A与野生型S.cerevisiaeW303-1A在甘油含量上相差不大,均只有4mmol/L左右。
Glycerol is an ideal osmotolerant medium. Two genes of GPD and GPP encoding glycerol 3- phosphate dehydrogenase and glycerol 3-phosphate phosphatase, key enzymes in glycerol synthesis, were cloned from Candida glycerolgenesis WL2002-5 using PCR method. In Escherichia coli JM109, enough GPD and GPP genes were amplified using T-vector, furthermore, expression plasmids pYX212-GPD containing GPD gene and pYX212-GPP containing GPP gene were constructed, which were introduced into Saccharomyces cerevisiae W303- 1A using LiAc transformation method successfully. Primary results showed that the biomass of pYX212-GPD/S. cerevisiae W303-1A was higher than those of pYX212-GPP/S, cerevisiae W303-1A and the wild type during the fermentation. After 72h fermentation, introduction of GPD gene could increase glycerol production to about 12mmoL/L, while GPP gene had little effect on glycerol production （only 4mmoL/L） in comparison with the wild type.
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2006年第1期38-41,共4页
China Biotechnology
基金
国家自然科学基金资助项目(30300027)
江南大学工业生物技术教育部重点实验室开放基金资助项目(KLIB-KF200507)
江南大学预研基金资助项目(0004402)
