JUN 14, 2022 9:00 AM PDT

Improving protein expression in E. coli B-strains using rapid genome engineering

Sponsored by: Inscripta
Speaker

Abstract
Date:  June 14, 2022
Time: 9:00am (PDT),  12:00pm (EDT), 6:00pm (CEST)
 
 
E. coli B-strains, such as BL21, and their derivatives are often used for protein expression due to their optimized amino acid production, ATP utilization, lon and ompT protease deficiency, and reduced acetate accumulation. However, optimizing B-strains for increased protein production using modern approaches to genome engineering has not been reported.  Here we apply CRISPR-based, high-throughput genome-wide and pathway-specific editing to improve the expression of an enzyme. With shallow sampling of the genome-edited libraries, we identify variants with greater than 2-fold improvement in active protein expression. This work demonstrates how rapid genome engineering can be applied to significantly enhance protein expression in E. coli.
 
 
Learning objectives:
  • Demonstrate genome-wide and pathway-specific editing in E. coli B-strains
  • Employ shallow sampling to find hits in the library with greater than 2-fold protein expression improvements
  • Identify protein expression hits in high throughput using automation strategies
 
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