Terpenoid production in microbes is mainly limited by the flux from intermediates of central metabolism (acetyl-CoA for MVA pathway or pyruvate and glyceraldehyde-3-phosphate for MEP pathway) to the substrates of TPSs (GPP, FPP, or GGPP) . The introduction and engineering of the biosynthetic MVA pathway in E. coli have improved the production of terpenoids such as amorphadiene , carotenoids [19, 21], coenzyme Q10 , and farnesol  by efficiently supplying IPP and DMAPP. Based on these studies, we exploited the biosynthetic MVA pathway for the biosynthesis of (−)-α-bisabolol in E. coli. To increase carbon flux towards TPS substrates, a plasmid (pSSN12Didi) containing the lower MVA pathway genes (Fig. 1b), leading from MVA to IPP and DMAPP , was introduced into E. coli DH5α expressing the MrBBS. MVA is an exogenous substrate for E. coli, so supplemented MVA can only be converted to IPP and DMAPP by the engineered MVA lower pathway in E. coli. Different concentrations of MVA, ranging from 0 to 10 mM, were examined to determine whether MVA availability limits (−)-α-bisabolol production in LB medium. (−)-α-bisabolol production increased as the concentration of the supplied MVA increased (Fig. 4a) and cell growth was not significantly inhibited (Fig. 4b). With the addition of 10 mM MVA, (−)-α-bisabolol production was 10.5 mg/L, approximately 3.5-fold higher than that obtained without the addition of MVA (3 mg/L, Fig. 4a). Expression of the lower MVA pathway genes in E. coli improved (−)-α-bisabolol production with the addition of MVA. However, MVA is not an economically viable substrate. To achieve (−)-α-bisabolol production from cost-effective renewable resources such as glycerol, a major byproduct of the biodiesel industry, we constructed a plasmid pSNA-MrBBS (Fig. 1b). pSNA-MrBBS harbors the complete biosynthetic MVA pathway genes, including the MrBBS gene, and enables production of IPP and DMAPP in the absence of MVA. A total of 12.8 mg/L of (−)-α-bisabolol was produced from glycerol in E. coli DH5α with pSNA-MrBBS grown in LB (Fig. 5a). This is a slightly higher concentration than that produced in DH5α containing only MrBBS and lower MVA pathway genes with the addition of 10 mM MVA (10.5 mg/L). Overall, the expression of biosynthetic MVA pathway genes significantly improved (−)-α-bisabolol production in E. coli.Fig. 4The effect of mevalonate concentration on (−)-α-bisabolol production (a) and final OD600 (b) from engineered E. coli harboring pTSN-MrBBS and pSSN12Didi. Cells were grown at 30 °C in LB medium supplemented with different concentrations of mevalonate and 20% (v/v) n-dodecane for 48 h. Data represent averages from three replicate cultures; error bars show SD Fig. 5Optimal conditions for (−)-α-bisabolol production in E. coli expressing MrBBS and all MVA pathway genes. E. coli cells harboring pSNA-MrBBS were grown for 48 h at 30 °C in the presence of 20% (v/v) n-dodecane with the following variations; a Medium (M9 supplemented with 4 g/L glucose, LB, 2xYT, TB) and glycerol concentration; b IPTG concentration. When OD600 reached 0.5, different concentrations of IPTG were added to the E. coli cells grown on TB medium supplemented with 10 g/L glycerol; c pH. Different initial pHs were tested to find an optimal pH for (−)-α-bisabolol production. E. coli cells were cultivated in TB medium containing 10 g/L glycerol in the absence of IPTG; d E. coli strains. E. coli cells were grown on TB medium, pH 7.0 with 10 g/L glycerol and 20% (v/v) n-dodecane in the absence of IPTG. Data represent averages from three replicate cultures and error bars show SD