Inactivation of an aldehyde/alcohol dehydrogenase gene fromClostridium acetobutylicum ATCC 824

Abstract
A nonreplicative plasmid containing an internalaad gene fragment has been integrated into the chromosome ofClostridium acetobutylicum ATCC 824. Transformation was accomplished by electroporation with relatively high concentrations of methylated plasmid DNA. Southern hybridization experiments revealed that integration occurred by single crossover homologous recombination inactivating theaad gene. Integrants were relatively stable after 25 generations. Inactivation of theaad gene drastically reduced solvent production. This result suggests that aldehyde/alcohol dehydrogenase (AAD) plays a important role in butanol production.