SWITCH: a dynamic CRISPR tool for genome engineering and metabolic pathway control for cell factory construction in Saccharomyces cerevisiae

Microbial Cell Factories

Research

Katherina García Vanegas¹  Uffe Hasbro Mortensen¹  Beata Joanna Lehka² 

[1] Department of Biotechnology and Biomedicine, Technical University of Denmark, Søltofts Plads, Building 223, Room 208, 2800, Kgs. Lyngby, Copenhagen, Denmark;Department of Science and Environment, Roskilde University, Universitetsvej 1, 4000, Roskilde, Denmark;

关键词: CRISPR tool;    Genome engineering;    Metabolic pathway control;    Cell factory;    Saccharomyces cerevisiae;   

DOI  :  10.1186/s12934-017-0632-x

 received in 2016-10-05, accepted in 2017-01-20,  发布年份 2017

来源: Springer

【 摘 要 】

BackgroundThe yeast Saccharomyces cerevisiae is increasingly used as a cell factory. However, cell factory construction time is a major obstacle towards using yeast for bio-production. Hence, tools to speed up cell factory construction are desirable.ResultsIn this study, we have developed a new Cas9/dCas9 based system, SWITCH, which allows Saccharomyces cerevisiae strains to iteratively alternate between a genetic engineering state and a pathway control state. Since Cas9 induced recombination events are crucial for SWITCH efficiency, we first developed a technique TAPE, which we have successfully used to address protospacer efficiency. As proof of concept of the use of SWITCH in cell factory construction, we have exploited the genetic engineering state of a SWITCH strain to insert the five genes necessary for naringenin production. Next, the naringenin cell factory was switched to the pathway control state where production was optimized by downregulating an essential gene TSC13, hence, reducing formation of a byproduct.ConclusionsWe have successfully integrated two CRISPR tools, one for genetic engineering and one for pathway control, into one system and successfully used it for cell factory construction.

【 授权许可】

CC BY   
© The Author(s) 2017