A rapid and efficient strategy for combinatorial repression of multiple genes in Escherichia coli.

ABSTRACT:

Background

The regulation of multiple gene expression is pivotal for metabolic engineering. Although CRISPR interference (CRISPRi) has been extensively utilized for multi-gene regulation, the construction of numerous single-guide RNA (sgRNA) expression plasmids for combinatorial regulation remains a significant challenge.

Results

In this study, we developed a combinatorial repression system for multiple genes by optimizing the expression of multi-sgRNA with various inducible promoters in Escherichia coli. We designed a modified Golden Gate Assembly method to rapidly construct the sgRNA expression plasmid p3gRNA-LTA. By optimizing both the promoter and the sgRNA handle sequence, we substantially mitigated undesired repression caused by the leaky expression of sgRNA. This meth

SUBMITTER: Zheng Y 

PROVIDER: S-EPMC11951683 | biostudies-literature | 2025 Mar

REPOSITORIES: biostudies-literature