Construction and characterization of shuttle vectors for succinic acid-producing rumen bacteriaopen access
- Authors
- Jang, Yu-Sin; Jung, Young Ryul; Lee, Sang Yup; Kim, Ji Mahn; Lee, Jeong Wook; Oh, Doo-Byoung; Kang, Hyun Ah; Kwon, Ohsuk; Jang, Seh Hee; Song, Hyohak; Lee, Sang Jun; Kang, Kyu Young
- Issue Date
- Sep-2007
- Publisher
- AMER SOC MICROBIOLOGY
- Citation
- APPLIED AND ENVIRONMENTAL MICROBIOLOGY, v.73, no.17, pp 5411 - 5420
- Pages
- 10
- Journal Title
- APPLIED AND ENVIRONMENTAL MICROBIOLOGY
- Volume
- 73
- Number
- 17
- Start Page
- 5411
- End Page
- 5420
- URI
- https://scholarworks.bwise.kr/cau/handle/2019.sw.cau/57007
- DOI
- 10.1128/AEM.01382-07
- ISSN
- 0099-2240
1098-5336
- Abstract
- Shuttle vectors carrying the origins of replication that function in Escherichia coli and two capnophilic rumen bacteria, Mannheimia succiniciproducens and Actinobacillus succinogenes, were constructed. These vectors were found to be present at ca. 10 copies per cell. They were found to be stably maintained in rumen bacteria during the serial subcultures in the absence of antibiotic pressure for 216 generations. By optimizing the electroporation condition, the transformation efficiencies of 3.0 X 10(6) and 7.1 X 10(6) transformants/mu g DNA were obtained with M. succiniciproducens and A. succinogenes, respectively. A 1.7-kb minimal replicon was identified that consists of the rep gene, four iterons, A+T-rich regions, and a dnaA box. It was found that the shuttle vector replicates via the theta mode, which was confirmed by sequence analysis and Southern hybridization. These shuttle vectors were found to be suitable as expression vectors as the homologous fumC gene encoding fumarase and the heterologous genes encoding green fluorescence protein and red fluorescence protein could be expressed successfully. Thus, the shuttle vectors developed in this study should be useful for genetic and metabolic engineering of succinic acid-producing rumen bacteria.
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