The invention belongs to the technical field of
biosynthesis, and particularly relates to a method for regulating and controlling
stevioside production by improving the
utilization rate of a
carbon source. According to the invention, a high-efficiency
stevioside biosynthesis system is constructed through systematic
engineering modification: in the aspect of
chassis cell optimization, transcription factors (AtWRKY18, AtWRKY40 and AtMYC2) for regulating and controlling MEP pathway and
key genes (dxs and / or dxr) of the MEP pathway are over-expressed, so that supply of
terpene precursors is enhanced; and a
heterologous MVA pathway (over-expression T7-atoB-mvaS-mvaE-mvK / mvKmut-pmK-mvaD-idi) with a
modular design is introduced, so that sufficient supply of the
terpenoid precursor is realized. Meanwhile, by knocking out related genes and bypass
metabolism genes (mgsA, iclR, menA or adhE) synthesized by
organic acid, a glucose transport and utilization
system (fnr, fadR, ppc, zwf or pgi is knocked out, and ACS, fbaA, tpiA, glf, galP or ptsG is overexpressed, so that the
utilization rate of a
carbon source is further improved, and the production capacity of
stevioside is remarkably improved.