Cell-free Directed Evolution of a Protease in Microdroplets at Ultrahigh Throughput.

ABSTRACT: Compartmentalization of single genes in water-in-oil emulsion droplets is a powerful approach to create millions of reactors for enzyme library selections. When these droplets are formed at ultrahigh throughput in microfluidic devices, their perfect monodispersity allows quantitative enzyme assays with a high precision readout. However, despite its potential for high quality cell-free screening experiments, previous demonstrations of enrichment have never been successfully followed up by actual enzyme library selections in monodisperse microfluidic droplets. Here we develop a three-step workflow separating three previously incompatible steps that thus far could not be carried out at once: first droplet-compartmentalized DNA is amplified by rolling circle amplification; only after completio

SUBMITTER: Holstein JM 

PROVIDER: S-EPMC7901014 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature