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In another study, Surzycki et al. (2007) developed an inducible system for chloroplast gene expression in Chlamydomonas. This system enabled the PSII activity to be turned off and thereby anaerobic conditions could be created when needed. Under the dark cycle, O2, which accumulated during photosynthesis, could be consumed by respiration. Furthermore, PSII could be reactivated by replenishing a minimum amount of sulfur. By repeating the dark activation–light production cycles, H2 production in the mutant Chlamydomonas was prolonged by up to 15 days.

2012. Sustained H-2 production in a Chlamydomonas reinhardtii D1 protein mutant. J. Biotechnol. 157 (4), 613–619. , 2012. Electron transfer in nitrogenase catalysis. Curr. Opin. Chem. Biol. 16 (1–2), 19–25. , 2012. Engineering of formate-hydrogen lyase gene cluster for improved hydrogen production in Escherichia coli. Int. J. Hydrogen Energy 37 (20), 15045–15051. , 2009. Recombineering: A homologous recombinationbased method of genetic engineering. Nat. Protoc. 4, 206–223. , 2011. Bioreactor and process design for biohydrogen production.

Improving photosynthesis and metabolic networks for the competitive production of phototroph-derived biofuels. Curr. Opin. Biotechnol. 23 (3), 290–297. , 2011. Microbial fuel cells come of age. J. Chem. Technol. Biotechnol. 86, 625–632. , 2009. Novel biological insights through metabolomics and C-13-flux analysis. Curr. Opin. Microbiol. 12, 553–558. C H A P T E R 3 Metabolic Engineering of Microorganisms for Biohydrogen Production You-Kwan Oh*1, Subramanian Mohan Raj†1, Gyoo Yeol Jung{, Sunghoon Park} * Clean Fuel Department, Korea Institute of Energy Research, Daejeon, Korea Center for Research and Development, PRIST University, Thanjavur, India { Department of Chemical Engineering & I-Bio Program, Pohang University of Science and Technology, Pohang, Korea } Department of Chemical and Biomolecular Engineering, Pusan National University, Busan, Korea † INTRODUCTION Hydrogen (H2) is a nonpolluting renewable fuel.

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