To genetically engineer Escherichia coli for the heterologous biosynthesis of triterpenoid, ambrein, the main bioactive component of ambergris, by constituting a novel squalene-derived ambrein biosynthetic pathway in E. coli. The ScERG9 gene encoding the squalene synthase (SS) was integrated into the E. coli genome to generate a squalene-producing strain that supplied the central precursor squalene for the formation of cyclic triterpenoids. The mutated squalene-hopene synthase (D377C SHC) and the tetraprenyl-beta-curcumene cyclase (BmeTC) were co-expressed with SS to construct a novel ambrein biosynthetic pathway in E. coli. Ambrein was produced at 2.6 mg l(-1). An E. coli chassis for ambrein production was constructed by combining the squalene synthesis module with the downstream cyclization module.