원문정보
초록
영어
Renewable energy production using bio-process has been considered to be a promising alternative for chemical conversion processes due to the depletion of fossil fuels. Genetic manipulation for yield enhancement and metabolite diversity also has been tried in the bio-process. Gram-positive bacteria such as acetogens which utilize synthesis gas have been used as biocatalysts; however, it is difficult to apply genetic tools for Gram-positive bacteria. The aim of this research is the construction of manipulated acetogen to increase the production of specific biochemicals. In this study, we applied the gene transformation technique to Eubacterium limosum KIST612 which was isolated from anaerobic digestor as strict anaerobic and acetogen. pKD46 designed for lambda recombinase was transformed into these bacteria. Although pKD46 was successfully transferred to E. imosum KIST612, the plasmids were not well recovered as the transformants. As an advanced approach, the homologous recombination was applied to knockout a target gene of genomic sequenced E. limosum KIST612. To make recombinants enhancing the acetate production, butyryl-CoA dehydrogenase encoding gene (bcd) was selected for the knockout. The shuttle vectors such as pBR322 and pJIR418 were used in this study. The homologous sequences of bcd gene were introduced to these vectors as an insert by the cloning method. After this, pBR322::bcd and pJIR418::bcd were in-vivo methylated via transformation with three different types of methylation pattern in Escherichiacoli strains and they will be introduced to E. limosum KIST612 by electrotransformation.