Dissertation > Industrial Technology > Chemical Industry > Basic inorganic chemical industry > Industrial gases > Hydrogen

Construction of Klebsiella Oxytoca HP1 Mutants by Homologous Recombination to Enhance Hydrogen Evolution

Author ZhuJunBo
Tutor XuFangCheng
School Xiamen University
Course Biochemical Engineering
Keywords Klebsiella oxytoca HP1 adhE gene LDHa gene Biohydrogen Gene insertion inactivation
CLC TQ116.2
Type Master's thesis
Year 2007
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Hydrogen bacterial hydrogen production capacity is not high biological hydrogen production technology is to limit the development of important factors . By means of the use of molecular biology to transform bacteria , is to improve the efficiency of the important biological hydrogen production method . Ethanol and lactic acid are Klebsiella Klebsiella oxytoca HP1 main anaerobic fermentation by-product H2 , cut useless byproducts increased H2 production rate and substrate conversion . In this study, K. oxytoca HP1 as a receptor , homologous arms was constructed integration vector pTA-Str and pLDH-Tr. Platform using endogenous double exchange principle , carried on the plasmid pMHE6 glucosaminidase adenosine transferase gene (Aminoglycoside-3'-adenyltransferase, aadA) exogenous expression cassette sequence ( containing a T7 promoter , aadA coding frame , T4 terminator, ) inserted into the recipient strain designated chromosome alcohol dehydrogenase ( containing acetaldehyde dehydrogenase and alcohol dehydrogenase ) gene adhE 868 ~ 869 zones (adhE start coding frame of 1 ) loci . And on this basis plasmid pTA-Str K. oxytoca HP1 adhE mediated insertion inactivation obtained with adhE- streptomycin resistant mutant . Using a similar method to carry on the pBR322 tetracycline resistance gene (tetracycline resistance protein, tet) exogenous expression cassette sequences ( including P1, P2 promoter , tet coding frame ) inserted into the recipient strain designated chromosome lactate dehydrogenase gene LDHa 518 ~ 519 zones (LDHa start coding frame of 1 ) loci . Based on this plasmid pLDH-Tr -mediated insertion of K. oxytoca HP1 LDHa inactivation obtained with tetracycline LDHa- resistant mutants . Experimental results show that the fermentation of glucose , the same fermentation conditions , adhE- mutant strain than in the wild hydrogen production increased 16.07% , ethanol production fell by 70.47%; LDHa- mutant strain than in the wild hydrogen production increased 20.35% . This study constructed Klebsiella oxytoca HP1 mutant adhE- and LDHa-, further research will lay the foundation for efficient hydrogen production system .

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