20.109(S22):Module 2
Contents
Module 2: metabolic engineering
Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)...
In this module you will use a modified CRISPR system to modulate transcription in E. coli. Specifically, you will use a CRISPR-based interference system to target a gene within the fermentation pathway of E. coli such that either ethanol or acetate production is increased.
Research goal: Increase the yield of commercially valuable byproducts in E.coli using CRIPSRi technology to target genes involved in mixed-acid fermentation pathway.
Lab links: day to day
M2D1: Complete in-silico cloning of pdCas9 expression plasmid
M2D2: Design gRNA sequence for CRIPSRi system
M2D3: Clone psgRNA expression plasmid
M2D4: Confirm gRNA sequence in psgRNA expression plasmid
M2D5: Prepare for induction of CRISPRi system
M2D6: Complete CRISPRi experiment and measure fermentation products
M2D7: Examine features in gRNA-targeted genomic sequences
M2D8: Organize figures and outline text for Research article
Major assignments
Journal Club presentation
Research article
References
- CRISPR interference (CRISPRi) for sequence-specific control of gene expression. Nature Methods. (2013) 8: 2180-2196.
- Metabolic engineering of Escherichia coli for production of mixed-acid fermentation end products. Frontiers in Bioengineering and Biotechnology. (2014) 2:1-12.
