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Jian-zhong Xu

http://orcid.org/0000-0001-6555-6059

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Journal of Zhejiang University SCIENCE B 2016 Vol.17 No.2 P.83-99

http://doi.org/10.1631/jzus.B1500187


Strategies used for genetically modifying bacterial genome: site-directed mutagenesis, gene inactivation, and gene over-expression


Author(s):  Jian-zhong Xu, Wei-guo Zhang

Affiliation(s):  The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi 214122, China

Corresponding email(s):   xujz126@126.com, zhangwg168@126.com

Key Words:  Escherichia coli, Corynebacterium glutamicum, DNA manipulation, Site-directed mutagenesis, Gene inactivation, Gene over-expression


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Jian-zhong Xu, Wei-guo Zhang. Strategies used for genetically modifying bacterial genome: site-directed mutagenesis, gene inactivation, and gene over-expression[J]. Journal of Zhejiang University Science B, 2016, 17(2): 83-99.

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T1 - Strategies used for genetically modifying bacterial genome: site-directed mutagenesis, gene inactivation, and gene over-expression
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DOI - 10.1631/jzus.B1500187


Abstract: 
With the availability of the whole genome sequence of Escherichia coli or Corynebacterium glutamicum, strategies for directed DNA manipulation have developed rapidly. DNA manipulation plays an important role in understanding the function of genes and in constructing novel engineering bacteria according to requirement. DNA manipulation involves modifying the autologous genes and expressing the heterogenous genes. Two alternative approaches, using electroporation linear DNA or recombinant suicide plasmid, allow a wide variety of DNA manipulation. However, the over-expression of the desired gene is generally executed via plasmid-mediation. The current review summarizes the common strategies used for genetically modifying E. coli and C. glutamicum genomes, and discusses the technical problem of multi-layered DNA manipulation. Strategies for gene over-expression via integrating into genome are proposed. This review is intended to be an accessible introduction to DNA manipulation within the bacterial genome for novices and a source of the latest experimental information for experienced investigators.

遗传改造细菌基因组的策略:基因定点突变、基因失活和基因过表达

概要:该综述较为全面地概述了当前针对大肠杆菌和谷氨酸棒杆菌基因组遗传改造的各个方法的具体流程、应用范围、注意事项以及其新颖之处,比较了针对基因定点突变、基因失活和基因过表达的各个方法所存在的优缺点,同时简单地介绍了利用质粒介导基因过表达所存在的问题。此外,还介绍了四种引物设计软件,并简单分析了它们的应用范围。为拟计划开展分子生物学实验的新手对关于细菌基因组遗传改造方法做了可靠的介绍,同时也为已进行相关实验的实验员提供关于基因定点突变、基因失活和基因过表达的最新信息。
关键词:大肠杆菌;谷氨酸棒杆菌;DNA操作方法;定点突变;基因失活;基因过表达

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