Investigation on the Effects of 5’- and 3’-Untranslated Regions on Recombinant Hydrogenase Activity in the Chlamydomonas Reinhardtii Chloroplast

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Abstract The availability of a Chlamydomonas Reinhardtii strain without a nuclear hydrogenase gene has opened the door for exploration of the Hyd gene in the chloroplast. Biolistics and homologous recombination provide a well understood tool for integrating recombinant genetic material into the C. Reinhardtii chloroplast. The effects of different 5’- and 3’-untranslated regions on hydrogen production catalyzed by a eukaryotic hydrogenase will be measured. 1 Introduction With the reduced stability of fossil fuel supplies and prices, interest in transportation fuels based on biological sources has been renewed. While terrestrial sources (e.g. corn ethanol and soybean biodiesel) currently replace a small fraction of fossil fuels, these fuel sources compete with agriculture for food. Aquatic phototrophs (algae and seaweed) are able to flourish in conditions unsuitable for agriculture, including saltwater, and may be easier to process into a usable fuel product.[12, 3, 6] So far, algae have been investigated for the production of lipids for biodiesel, starches for ethanol production, and direct production of hydrogen gas.[9, 2] The natural propensity of certain algae to produce hydrogen gas under anaerobic condi- tions has lead to interest in optimizing the metabolic processes involved, such that algae could be made to efficiently convert water into hydrogen gas[9, 2] (in contrast to photovoltaic sys- tems, which involve expensive solar panels and inefficient electrolytic processes). The purpose of this project is to study the effects of untranslated DNA regions related to the hydrogenase enzyme on hydrogen production in Chlamydomonas Reinhardtii. 2 Background 2.1 Chlamydomonas Reinhardtii Chlamydomonas Reinhardtii has become a m... ... middle of paper ... ...hloro- plast transformation in chlamydomonas reinhardtii. Methods Mol Biol 274 (2004), 301– 307. [14] Rasala, B. A., Muto, M., Lee, P. A., Jager, M., Cardoso, R. M., Behnke, C. A., Kirk, P., Hokanson, C. A., Crea, R., Mendez, M., and Mayfield, S. P. Production of therapeutic proteins in algae, analysis of expression of seven human proteins in the chloroplast of chlamydomonas reinhardtii. Plant Biotechnol J (Mar 2010). [15] Remacle, C., Cardol, P., Coosemans, N., Gaisne, M., and Bonnefoy, N. High- efficiency biolistic transformation of chlamydomonas mitochondria can be used to insert mutations in complex i genes. Proc Natl Acad Sci U S A 103, 12 (Mar 2006), 4771–4776. 4 [16] Ru ̈hle, T., Hemschemeier, A., Melis, A., and Happe, T. A novel screening pro- tocol for the isolation of hydrogen producing chlamydomonas reinhardtii strains. BMC Plant Biol 8 (2008), 107–107.

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