Multiplex Genome Editing by Natural Transformation (MuGENT) for Synthetic Biology in Vibrio natriegens.
Multiplex Genome Editing by Natural Transformation (MuGENT) for Synthetic Biology in Vibrio natriegens.
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DOI：
10.1021/acssynbio.7b00116
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发表时间：
2017-09-15
影响因子：
4.7
通讯作者：
Dalia AB
中科院分区：
文献类型：
--
作者：
Dalia TN;Hayes CA;Stolyar S;Marx CJ;McKinlay JB;Dalia AB
产钠弧菌最近成为分子生物学和生物技术中大肠杆菌的替代品，但低效的遗传工具阻碍了其发展。在这里，我们揭示了如何在产钠弧菌中诱导天然感受态，并描述了通过自然转化进行多重基因组编辑（MuGENT）的方法。MuGENT在前所未有的时间尺度上以高效率促进多个基因组编辑的整合。此外，该方法允许产生高度复杂的突变体群体，其可用于代谢工程努力。作为概念验证，我们试图通过MuGENT靶向参与PHB生物合成的9个基因的表达，来提高产钠弧菌中增值化学品聚β-羟基丁酸酯（PHB）的产量。在1周内，我们分离出编辑的菌株，其产生的PHB比亲本分离株多约100倍，比合理设计的菌株多约3.3倍。因此，这里描述的方法应该扩展该物种的实用性，用于各种学术和工业应用。
Vibrio natriegens has recently emerged as an alternative to Escherichia coli for molecular biology and biotechnology, but low-efficiency genetic tools hamper its development. Here, we uncover how to induce natural competence in V. natriegens and describe methods for multiplex genome editing by natural transformation (MuGENT). MuGENT promotes integration of multiple genome edits at high-efficiency on unprecedented timescales. Also, this method allows for generating highly complex mutant populations, which can be exploited for metabolic engineering efforts. As a proof-of-concept, we attempted to enhance production of the value added chemical poly-β-hydroxybutyrate (PHB) in V. natriegens by targeting the expression of nine genes involved in PHB biosynthesis via MuGENT. Within 1 week, we isolated edited strains that produced ~100 times more PHB than the parent isolate and ~3.3 times more than a rationally designed strain. Thus, the methods described here should extend the utility of this species for diverse academic and industrial applications.
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影响因子：
3.2
作者：
Dalia, Ankur B.;Lazinski, David W.;Camilli, Andrew
通讯作者：
Camilli, Andrew
影响因子：
4.4
作者：
Gulig, Paul A.;Tucker, Matthew S.;Brown, Roslyn N.
通讯作者：
Brown, Roslyn N.
影响因子：
4.4
作者：
KARR, DB;WATERS, JK;EMERICH, DW
通讯作者：
EMERICH, DW
影响因子：
5.1
作者：
Pollack-Berti A;Wollenberg MS;Ruby EG
通讯作者：
Ruby EG
影响因子：
64.8
作者：
Wang HH;Isaacs FJ;Carr PA;Sun ZZ;Xu G;Forest CR;Church GM
通讯作者：
Church GM