An ultrahigh‐throughput screening platform based on flow cytometric droplet sorting for mining novel enzymes from metagenomic libraries. (10th October 2020)

• Record Type: Journal Article
• Authors: Ma, Fuqiang; Guo, Tianjie; Zhang, Yifan; Bai, Xue; Li, Changlong; Lu, Zelin; Deng, Xi; Li, Daixi; Kurabayashi, Katsuo; Yang, Guang‐yu
• Other Names: Liu Shuang‐Jiang guestEditor.; Wu Xiao‐Lei guestEditor.; Huang Wei guestEditor.; Zhang Yu‐Zhong guestEditor.; Wang Fengping guestEditor.
• Is Part Of: Environmental microbiology. Volume 23:Number 2(2021)
• Page Start: 996
• Page End: 1008
• Publication Date: 2020-10-10
• Subjects: Microbial ecology -- Periodicals; Environmental Microbiology -- Periodicals; 579.17
• DOI: 10.1111/1462-2920.15257
• Languages: English
• ISSNs: 1462-2912

Abstract

Summary: Uncultivable microbial communities provide enormous reservoirs of enzymes, but their experimental identification by functional metagenomics is challenging, mainly due to the difficulty of screening enormous metagenomic libraries. Here, we propose a reliable and convenient ultrahigh‐throughput screening platform based on flow cytometric droplet sorting (FCDS). The FCDS platform employs water‐in‐oil‐in‐water double emulsion droplets serving as single‐cell enzymatic micro‐reactors and a commercially available flow cytometer, and it can efficiently isolate novel biocatalysts from metagenomic libraries by processing single cells as many as 10 8 per day. We demonstrated the power of this platform by screening a metagenomic library constructed from domestic running water samples. The FCDS assay screened 30 million micro‐reactors in only 1 h, yielding a collection of esterase genes. Among these positive hits, Est WY was identified as a novel esterase with high catalytic efficiency and distinct evolutionary origin from other lipolytic enzymes. Our study manifests that the FCDS platform is a robust tool for functional metagenomics, with the potential to significantly improve the efficiency of exploring novel enzymes from nature.