大肠杆菌甘油激酶基因(glpK)和甘油脱氢酶基因(gldA)的敲除及其对甘油产量的影响

利用Red重组系统构建了大肠杆菌JM109甘油激酶基因(glpK)和甘油脱氢酶基因(gldA)缺失的双突变菌株JM109B,然后将表达酿酒酵母3-磷酸甘油脱氢酶基因(GPD1)和3-磷酸甘油酯酶基因(HOR2)的质粒pSE-gpd1-hor2转化到JM109B突变菌株中,在含1%葡萄糖的摇瓶发酵培养基中37℃发酵24 h,甘油的最高产量为5.61 g/L,是原始菌株JM109/pSE-gpd1-hor2甘油产量的1.59倍;在30 L发酵罐中发酵28 h,甘油的最高产量为103.12 g/L,是原始菌株JM109/pSE-gpd1-hor2甘油产量的1.59倍,是原始菌株BL21/pSE-gpd1-hor2甘油产量的1.41倍,葡萄糖转化率为50.39%。

Disruption of glpK and gldA genes in E.coli JM109 and effects on yield of glycerol

By using Red recombination system, E. coli JM109 mutational strain with genes glpK and gldA disruption was constructed. The mutational strain was named E. coli JM109B. Then, the recombinant plasmid pSE-gpdl-hor2, which expressed 3-phosphate dehydrogenase and glycerol 3-phosphatase from Saccharornyces cerevisiae, was transformed into E. coli JM109B. And the recombinant microorganism was incubated in minimal media with 1% glucose shake-flasks at 37 12 with vigorous shaking for 24 h. Its maximal concentration of glycerol was 5.61 g/L, which was 1.59 times than that of E. coli recombinant strain JM109/ pSE-gpdl-hor2. The recombinant strain was fermented at 37 12 for 28 h. The maximal concentration of glycerol was 103.12 g/L, which was 1.59 times than that d E. coli JM109/pSE-gpdl- hor2 and was 1.41 times than that of E. coli BI21/pSE-gpdl-hor2, respectively. The conversion rate of glucose to glycerol was 50.39 %.