The invention relates to a method for preparing
rebaudioside A by liquid
fermentation. The method includes: (1) transferring a
glycosyltransferase UGT76G1
gene to a pUC18
plasmid vector, transferringto DH5alpha
escherichia coli competent cells, inoculating into an LB medium, and culturing for 10-18 hours at 30-37 DEG C and 200-250 rpm; (2) inoculating into a seed tank according to an inoculationamount of 0.1-10% by volume, and culturing for 5-12 hours at 100-400 rpm, a ventilation ratio of 0.2-1 V / V. min and 30-37 DEG C; (3) inoculating into a
fermentation tank according to an
inoculation amount of 1-15% by volume, and culturing for 20-30 hours at 100-800 rpm, a ventilation ratio of 0.5-2 V / V. min and 30-37 DEG C; (4) adding an
inducer when
cell concentration OD600 reaches 20-60, whereinthe concentration is 0.05-1 mmol / L; (5) carrying out pressure
filtration, resuspension, high-pressure homogenization and crushing, and pressure
filtration to obtain a crude
enzyme liquid; and (6) mixing
stevioside,
uridine diphosphate glucose, a
phosphate buffer solution and the crude
enzyme liquid, and reacting at 25-40 DEG C for 12-48h. The method high
enzyme activity, high catalytic substrateconcentration, high conversion rate, easy control of the
fermentation process, simple steps and low fermentation cost.