We previously synthesized poly(3-hydroxybutyrate) [P(3HB)] in recombinant Corynebacterium glutamicum, a prominent producer of amino acids. In this study, a two-step cultivation was established for the dual production of glutamate and P(3HB) due to the differences in the optimal concentration of biotin. Glutamate was extracellularly produced first under the biotin-limited condition of 0.3 mu g/L. Production was then shifted to P(3HB) by addition of biotin to a total concentration of 9 mu g/L. The final products obtained were 18 g/L glutamate and 36 wt% of P(3HB). (c) 2008, The Society for Biotechnology, Japan. All rights reserved.