Biotechnology Bulletin ›› 2023, Vol. 39 ›› Issue (10): 93-106.doi: 10.13560/j.cnki.biotech.bull.1985.2023-0486

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Application of FACS Technology in the Directed Evolution of Enzyme

LIU Jin-sheng(), CHEN Zhen-ya(), HUO Yi-xin, GUO Shu-yuan()   

  1. School of Life Science, Beijing Institute of Technology, Beijing 100081
  • Received:2023-05-23 Online:2023-10-26 Published:2023-11-28
  • Contact: CHEN Zhen-ya, GUO Shu-yuan E-mail:ljs15536837748@163.com;chenzhenya@bit.edu.cn;guosy@bit.edu.cn

Abstract:

Flow cytometry(FCM)is a rapid and relatively quantitative multi-parameter analytical technique for analyzing a large number of cells at the single-cell level. The constructed fluorescence activated cell sorting(FACS)based on FCM can physically separate fluorescent-labeled single cells and has been widely used in the field of directed evolution of enzyme. Directed evolution has been proven to be an effective method for obtaining industrial enzymes with excellent properties, such as high enzyme activity, strong thermal stability and solvent resistance. To conduct directed evolution, mutant library firstly needs to be constructed by random mutagenesis or DNA recombination, and then be screened under manual selection pressure. Traditional screening methods such as plate and microplate methods have limitations of low throughput, high cost, large errors, and poor accuracy, and cannot achieve high-throughput screening of large mutant library. Compared with traditional screening methods, FACS has the advantages of high throughput, low cost, small error and high precision. In this article, we first summarized the development process of FCM and FACS, as well as the composition and classification of related instruments derived from them. Then, we discussed the application status and limitations of FACS in the directed evolution of enzyme. Finally, we summarized and prospected the development direction of FACS and its application in the field of directed evolution of enzyme.

Key words: flow cytometry, fluorescence-activated cell sorting, high-throughput screening