期刊
METABOLIC ENGINEERING
卷 54, 期 -, 页码 232-243出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymben.2019.04.007
关键词
High-throughput strain screening; Cross-feeding; Microbial co-culture; Droplet microfluidics; Biosensor; Signal amplification; Industrial microbiology; Metabolic engineering
资金
- USDA AFRI NIFA Fellowships Grant Program [2016-67011-24725]
- NIFA [2016-67011-24725, 810678] Funding Source: Federal RePORTER
Microbes can be engineered to synthesize a wide array of bioproducts, yet production phenotype evaluation remains a frequent bottleneck in the design-build-test cycle where strain development requires iterative rounds of library construction and testing. Here, we present Syntrophic Co-culture Amplification of Production phenotype (SnoCAP). Through a metabolic cross-feeding circuit, the production level of a target molecule is translated into highly distinguishable co-culture growth characteristics, which amplifies differences in production into highly distinguishable growth phenotypes. We demonstrate SnoCAP with the screening of Escherichia coil strains for production of two target molecules: 2-ketoisovalerate, a precursor of the drop-in biofuel isobutanol, and L-tryptophan. The dynamic range of the screening can be tuned by employing an inhibitory analog of the target molecule. Screening based on this framework requires compartmentalization of individual producers with the sensor strain. We explore three formats of implementation with increasing throughput capability: confinement in microliter plates (10(2)-10(4) assays/experiment), spatial separation on agar plates (10(4)-10(5) assays/experiment), and encapsulation in microdroplets (10(5)-10(7) assays/experiment). Using SnoCAP, we identified an efficient isobutanol production strain from a random mutagenesis library, reaching a final titer that is 5-fold higher than that of the parent strain. The framework can also be extended to screening for secondary metabolite production using a push-pull strategy. We expect that SnoCAP can be readily adapted to the screening of various microbial species, to improve production of a wide range of target molecules.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据