高产二羟基丙酮重组菌株的构建

Construction of a Recombinant Strain Producing High-yield Dihydroxyacetone

旨在构建一株过量表达编码膜系甘油脱氢酶的sld AB基因的重组菌株,以提高二羟基丙酮产量。以氧化葡萄糖酸杆菌ATCC621H的基因组DNA为模板,运用PCR方法扩增得到基因sld AB,并连接到p BBR1MCS-2质粒上,构建表达载体p BBR1MCS-2-sld AB。通过电转化将载体p BBR1MCS-2-sld AB转化进入氧化葡萄糖酸杆菌ATCC621H内,得到重组菌株GOX205。结果显示,重组菌株构建成功,其甘油脱氢酶的酶活力较之于出发菌株提高了26%。在甘油初始浓度100 g/L的甘油发酵培养基中,较之于出发菌株,GOX205的生长状况良好,发酵52 h时DHA浓度达到94.1 g/L,较之于出发菌株提高了19.7%,甘油残量降低了15.1 g/L。

This work aims to construct a recombinant strain containing gene sldAB over-expressing membrane-bound glycerol dehydrogenase for raising the yield of dihydroxyacetone （ DHA ） . Firstly, using genomic DNA of Gluconobacter oxydans ATCC621H as a template, sldAB amplified by PCR was ligated to the plasmid pBBR1MCS-2 and the expression vector pBBR1MCS-2-sldAB was constructed ; subsequently the plasmid pBBR1MCS-2-sldAB was transformed into the G. oxydans ATCC621H by electrotransfer, and the recombinant strain GOX205 was obtained. Results showed that the recombinant strain was constructed successfully, and the activity of glycerol dehydrogenase increased by 26% compared with the original strain. When initial concentration 100 g/L of glycerol as carbon source, the growth of GOX205 was significantly improved compared with the original strain, the final DHA concentration was 94.1 g/L and increased by 19.7%, and the residue of glycerol decreased 15.1 g/L. Therefore, this study provides a basis for further improving the biotransformation process of DHA production.