Arrayed CRISPRi library to suppress genes required for Schizosaccharomyces pombe viability

Area of Science:

• Molecular Biology
• Genetics
• Cell Biology

Background:

• Fission yeast (Schizosaccharomyces pombe) is a key model organism for eukaryotic biological processes.
• Understanding essential genes is crucial, but requires tools for conditional gene manipulation.
• Existing methods may not comprehensively target the approximately 1,200 essential genes in S. pombe.

Purpose of the Study:

• To create versatile CRISPR interference (CRISPRi) libraries for targeted gene knockdown in S. pombe.
• To cover a significant portion of essential genes for functional genomics studies.
• To facilitate the investigation of essential gene functions and their roles in cellular processes.

Main Methods:

• Construction of arrayed plasmid and strain libraries for S. pombe.
• Utilizing dCas9-mediated CRISPRi for targeted gene repression.
• High-throughput screening and validation of knockdown efficiency.

Main Results:

• Libraries encompass approximately 98% of essential genes in fission yeast.
• Efficient transcriptional repression, inhibiting cell proliferation in about 60% of targeted strains.
• Demonstrated utility through metabolic analyses, revealing pathway interactions.

Conclusions:

• The developed CRISPRi libraries provide a powerful resource for comprehensive functional genomics in S. pombe.
• These tools will advance the understanding of essential gene functions in eukaryotes.
• The libraries enable detailed investigation of metabolic flexibility and other biological processes.