This study investigates the enhancement of bioethanol production using a genetic engineering approach. The bioethanol-producing strain, E. aerogenes ATCC 29007, was engineered by deleting the D-lactate dehydrogenase (ldhA) gene to block the production of lactic acid. The Open-reading frame coding region of ldhA gene was replaced with a kanamycin cassette flanked by FLP recognition target sites by using a one-step method to inactivate chromosomal genes and primers designed to create in-frame deletions upon excision of the resistance cassette. The colony PCR was used to confirm the deleted gene. Glycerol, a useful byproduct in the biodiesel industry, was employed to convert into bioethanol, using engineered E. aerogenes SUMI014. Under optimal conditions of fermentation (34 °C, pH 7.5, 78 h), bioethanol production by the mutant strain was 34.54 g/L, 1.5 times greater than that produced by its wild type (13.09 g/L). Subsequent overexpression of alcohol dehydrogenase (adhE) gene in the mutant strain; increased the production of bioethanol up to 38.32 g/L. By the combination of gene deletion and overexpression, the bioethanol yield was 0.48 g/g when employing 80 g/L glycerol. Hence, a significant enhancement in ethanol production was observed. These results may provide valuable guidelines for further engineering bioethanol producers.
Bibliographical noteFunding Information:
This work was supported by the Advanced Biomass R&D Center (ABC-2011-0031360) of Global Frontier Project funded by the Ministry of Science, ICT and Future Planning of Korea and Basic Science Research Program through the National Research Foundation of Korea funded by the Ministry of Education (NRF-2013R1A1A2057887).
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- Alcohol dehydrogenase
- Enterobacter aerogenes ATCC29007
- Enterobacter aerogenes SUMI014
- Enterobacter aerogenes SUMI2008
- d-lactate dehydrogenase
ASJC Scopus subject areas
- Applied Microbiology and Biotechnology