High-Throughput Screening of L-Lactic Acid-Hyperproducing Bacillus coagulans Mutants Using an Automated Droplet Microfluidic Platform

Le résumé fourni par la source

Industrial biomanufacturing relies on microorganisms to efficiently synthesize target products under engineered conditions. However, wild-type strains often lack the capability to do so due to their native metabolic networks. Therefore, the rapid and accurate screening of high-performance strains from large-scale mutant libraries is a critical step in industrial microbial breeding. In recent years, droplet microfluidic technology has attracted significant attention for its high throughput, low consumption, and single-cell compartmentalization capabilities, leading to the development of numerous single-cell sorting systems. A high‑throughput screening protocol for industrial microbial strains using an automated droplet microfluidic platform is presented in this study. The integrated workflow comprises four key stages: construction of a mutant library via atmospheric and room-temperature plasma (ARTP) mutagenesis; high‑efficiency single‑cell encapsulation and controlled micro‑cultivation within picoliter‑scale droplets; precise pico‑injection of a fluorescent biosensor, followed by fluorescence‑activated droplet sorting (FADS) to isolate droplets containing high‑producing variants; and validation of the sorted candidate strains through microplate and shake‑flask fermentation. A complete and integrated screening workflow is intended as a standardized template, and its validation is demonstrated through the isolation of high‑yielding L-lactic acid‑producing mutants of B. coagulans as a specific example. This automated, high-throughput screening platform offers a powerful technical solution with significant potential to accelerate industrial breeding of high-performance strains.

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