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中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志
中国生物工程杂志  2019, Vol. 39 Issue (11): 113-122    DOI: 10.13523/j.cb.20191113
综述     
人工合成多样性突变文库研究进展*
王兆官,吴洋,齐浩
天津大学化工学院 系统生物工程教育部重点实验室 天津化学化工协同创新中心合成生物学平台 天津 300072
Research Progress in Synthetic Diverse Mutant Libraries
WANG Zhao-guan,WU Yang,QI Hao
School of Chemical Engineering and Technology, Key Laboratory of Systems Bioengineering of Ministry of Education,Syn Bio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering,Tianjin University, Tianjin 300072, China
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摘要：

突变文库的构建是定向进化研究过程中一个关键步骤,主要利用天然存在的系统或者人工合成的分子技术来产生多样性核酸分子文库,为制备和筛选具有一定特性的蛋白酶、多肽、人工抗体等提供庞大的遗传基因库,也可用于合成生物学中相关基因元件的研究与筛选,为目标生物制品的高效工业化生产提供动力。随着对突变文库构建技术研究的日益深入,各种文库构建策略相继被开发出来,并在生物能源、生物化工、生物医药、生物试剂和食品工业等方面得到了广泛的应用。然而,定向进化中的文库构建策略多有不同,各种突变文库构建技术的核心方法也在不断创新。主要介绍近年来实验室中人工合成多样性文库的前沿技术,并对文库构建技术在自动化和智能化方向的发展进行了展望。
关键词： 定向进化;  体外突变;  人工合成;  基因文库    
Abstract:

The construction of mutant library is a key step in the process of directed evolution. It mainly utilizes the system of natural or synthetic molecular technique to generate the diversity of the nucleic acid molecular libraries. It also provides large genetic gene pool for the preparation and screening of protease, polypeptide and artificial antibodies with specific properties. Furthermore, it can be used in the study and selection of genetic element in synthetic biology, providing power to highly industrialized production of biological products. With the deepening of the research on the construction technology of mutant libraries, various library construction strategies have been developed and widely applied in the fields of bioenergy, biochemistry, biomedicine, biological reagent and food industry. However, the strategies of library construction in the directed evolution are different, and the core methods of various mutant library construction technologies are constantly innovated. Here it mainly elucidates the leading technologies of synthesizing multiplex libraries for the laboratory in recent years, and prospects for the development of library construction technology in the direction of automation and intellectualization.
Key words: Directed    evolution    In    vitro    mutation    Artificial    synjournal    Gene    library
收稿日期: 2019-02-17 出版日期: 2019-12-17
ZTFLH:  Q819  
基金资助: * 国家自然科学基金(21476167、21778039)
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王兆官
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引用本文:

王兆官,吴洋,齐浩. 人工合成多样性突变文库研究进展*[J]. 中国生物工程杂志, 2019, 39(11): 113-122.

WANG Zhao-guan,WU Yang,QI Hao. Research Progress in Synthetic Diverse Mutant Libraries. China Biotechnology, 2019, 39(11): 113-122.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20191113    或    https://manu60.magtech.com.cn/biotech/CN/Y2019/V39/I11/113

图1  无装配的的突变文库构建策略
图2  几种突变文库装配策略
图3  几种基于单链DNA的突变文库构建策略
NameTarget of
mutation		Theoretic
library size		Modification
primers		EfficiencyReference
PFLF-MSDMLacZ(6) and (4)6.4 ×107No40%~80%[21]
In situ epPCRxynARandomNo50%~85%[32]
OD-SPMNeo (11)2.05×10145' phosphorylation100%[38]
POEPLacZ (8)3.2 ×106No100%[40]
TEDAphbCAB(3)8×103No90%[42]
OmniChangePhytase (5)3.2 ×1065'phosphorothioate100%[45]
Simple-MSSMeGFP (5)3.2 ×106No100%[48]
ICMGFP (2)400No94.5%[49]
MegAnnealscFvs200-750ntRandomNo37%~79%[55]
PFunkelBla (4)1.6 ×1055' phosphorylation70%[56]
Nicking MutagenesisBla (3)8×1035' phosphorylation50%[57]
Darwin AssemblyKOD (19)5.24×10245' phosphorylation98%~100%[58]
表1  部分突变技术介绍
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