Polyhydroxyalkanoates (PHAs) are biodegradable polyesters accumulated in a wide variety of microorganisms as intracellular carbon and energy storage compounds. Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) is one of the most valuable biopolymers because of its superior mechanical properties. Here, we developed a bioprocess utilizing recombinantBacillus megateriumstrain for PHBV over-production from glucose, without any precursor addition. PHA production was performed in a controlled bioreactor by batch and fed-batch modes using wild-typeB. megateriumand rec-B. megateriumcells overexpressing the nativephaCgene. The effect of oxygen transfer rate on biomass formation and PHA accumulation was also investigated, under different dissolved oxygen levels. Structural and thermal properties of PHA were characterized by GC-FID,H-1-NMR, TGA and DSC analyses. Significantly, the copolymer produced from glucose as the carbon source in rec-B. megateriumwas composed of 58 mol% of 3-hydroxyvalerate monomers. After 66 h, rec-B. megateriumcells in fed-batch fermentation with a pre-determined growth rate mu(0) = 0.1 h(-1)produced the highest CDW (7.7 g L-1) and PHA concentration (6.1 g L-1). Moreover, an exponential glucose feeding profile resulted in 2.2-fold increase in PHA yield compared to batch cultivation. Overall, this study paves the way to an enhanced biopolymer production process inB. megateriumcells, where the highest product yield on cell was obtained asY(P/X) = 0.8 g g(-1).