| Autor: |
Lee YR; National Biotechnology Policy Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 34141, Republic of Korea., Lee J; School of Earth Sciences and Environmental Engineering, Gwangju Institute of Science and Technology (GIST), Gwangju 61005, Republic of Korea., Hong S; Department of Bio-Environmental Chemistry, College of Agricultural and Life Sciences, Chungnam National University, Daejeon 34134, Republic of Korea., Lee SY; Gwangju Clean Energy Research Center, Korea Institute of Energy Research, Gwangju, 61003, Republic of Korea., Lee WH; Department of Bioenergy Science and Technology, Chonnam National University, Gwangju 61186, Republic of Korea., Koh M; Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan 46241, Republic of Korea., Chang IS; School of Earth Sciences and Environmental Engineering, Gwangju Institute of Science and Technology (GIST), Gwangju 61005, Republic of Korea., Lee S; Department of Bio-Environmental Chemistry, College of Agricultural and Life Sciences, Chungnam National University, Daejeon 34134, Republic of Korea. |
| Abstrakt: |
Rhodobacter sphaeroides is a strain capable of both photoautotrophic and chemoautotrophic growth, with various metabolic pathways that make it highly suitable for converting carbon dioxide into high value-added products. However, its low transformation efficiency has posed challenges for genetic and metabolic engineering of this strain. In this study, we aimed to increase the transformation efficiency of R. sphaeroides by deleting the rshI gene coding for an endogenous DNA restriction enzyme that inhibits. We evaluated the effects of growth conditions for making electrocompetent cells and optimized electroporation parameters to be a cuvette width of 0.1 cm, an electric field strength of 30 kV/cm, a resistance of 200 Ω, and a plasmid DNA amount of 0.5 μg, followed by a 24-h recovery period. As a result, we observed over 7,000 transformants per μg of DNA under the optimized electroporation conditions using the R. sphaeroides ΔrshI strain, which is approximately 10 times higher than that of wild-type R. sphaeroides under standard bacterial electroporation conditions. These findings are expected to enhance the application of R. sphaeroides in various industrial fields in the future. |
