In this study, we have engineered E. coli for de novo production of (−)-α-bisabolol for the first time. Introduction of the MVA pathway, (−)-α-bisabolol synthase from German chamomile, and FPP synthase turned E. coli into a microbial cell factory for the de novo production of (−)-α-bisabolol from the renewable carbon source, glycerol. Using Canola oil as an extraction solvent, the engineered E. coli strain produced 9.1 g/L of (−)-α-bisabolol from glycerol in a fed-batch fermentation system, and the specific (−)-α-bisabolol production and productivity reached 0.18 g/g DCW and 1.24 g/L/day, respectively. To the best of our knowledge, this is the first report demonstrating that a large amount of (−)-α-bisabolol could be produced by metabolically engineered E. coli. Although more work is needed to optimize the fermentation process, the strains developed in this study will serve as promising platform hosts for development of microbial production of (−)-α-bisabolol on a large scale. Moreover, a more environmentally friendly means to extract (−)-α-bisabolol, using vegetable oils, will reduce the production cost in an industrial process.