Earticle

초록

영어

Zymomonas mobilis ZM4 was an important gram-negative bacterium used for fuel ethanol production and biorefinary process. For a long time, pLOI193 has been used as the most efficient mobilizable plasmid for Z. mobilis strains; however, the limited cloning sites and relative big size (13.4kb) of this plasmid has limited its application in molecular manipulation. In this work, a new plasmid pHW20a as tool vector was constructed from pLOI193 for efficient transformation of Z. mobilis cells. The constructed vector contains an oriV for replication in gram-negative bacteria, one antibiotic resistant gene (tetracycline resistant gene, tcr), two mob genes (mob from RP4 and mob from RSF1010), and a screening marker lacZα for genes to be cloned into the multiple cloning sites by blue-white screening. The transconjugation efficiency of pHW20a was about two orders of magnitude greater than pLOI193 and about three orders of magnitude higher than pBBR1MCS derivative plasmid. The constructed pHW20a could be stably transferred in Z. mobilis ZM4 without selection pressure at least for 120
generations and the function of lacZα was also confirmed by blue-white screening with insertion of genes in the multiple cloning sites. Two examples of Z. mobilis ZM4 transformation were demonstrated by overexpressing a Z. mobilis ZM4 native gfo gene (encoding glucose-fructose oxidoreductase, GFOR) and expressing a heterologous mdh gene (encoding malate dehydrogenase, MDH) from E. coli BL21, respectively. The results indicated that pHW20a was a useful vector for transformation of Z. mobilis strains. A low level of ppc transcription was also observed during the expression of mdh under the control of ppc promoter. The low ppc transcription level was considered as another important factor responsible for the uncoupled growth of Z. mobilis ZM4, although a low ATP generation efficiency of ED pathway was traditionally regarded as the sole reason for the uncoupled growth. The comprehensive understanding of the uncoupled growth of ZM4 would be beneficial to the effective preparation of biocatalysts for the fuel ethanol and sorbitol production with a finely tuned strategy.

키워드

저자

• Hong-Wei Dong [ State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China. ]
• Jie Bao [ State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China. ] Corresponding authors
• Dewey D. Y. Ryu [ Biochemical Engineering Program, University of California, One Shields Ave, Davis, CA 95616, USA. ]
• Jian-Jiang Zhong [ State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China., Key Laboratory of Microbial Metabolism of Ministry of Education, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China. ] Corresponding authors

참고문헌

발행기관

간행물

표지보기 관심저널 등록

• 간행물명

  한국생물공학회 학술대회

• 간기

  반년간

• 수록기간

  1985~2013

• 십진분류

  KDC 476 DDC 576