CRISPR-Based Approaches for Gene Regulation in Non-Model Bacteria

Area of Science:

• Microbiology
• Molecular Biology
• Synthetic Biology

Background:

• CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa) are widely used for gene expression control in bacteria.
• These tools enable dynamic engineering of cellular metabolism, transcriptional circuitry, and genotype-phenotype relationships.
• While established in model bacteria like *E. coli*, their application is expanding to non-model species.

Purpose of the Study:

• To review the challenges and current status of CRISPRi/a tool development in diverse non-model bacteria.
• To summarize the application landscape of CRISPRi/a across different bacterial phyla.
• To identify barriers and suggest future directions for broader CRISPRi/a implementation.

Main Methods:

• Literature review of studies reporting CRISPRi/a in non-model bacteria.
• Analysis of application areas including tool development, gene function interrogation, metabolic engineering, and high-throughput screening.
• Categorization of reported studies across bacterial phyla.

Main Results:

• Over 190 recent examples of CRISPRi/a applications in non-model bacteria across eight phyla were identified.
• Most studies focused on tool development and gene function studies, with fewer applications in metabolic engineering or high-throughput screens.
• The majority of reported CRISPRi/a tools target common strains, indicating challenges in applying them to undomesticated bacteria.

Conclusions:

• CRISPRi/a tools show significant adoption in diverse non-model bacteria, demonstrating their versatility.
• Further development of efficient and generalizable methods is crucial for wider implementation, especially in undomesticated strains.
• Programmable CRISPR-based transcriptional control holds immense potential for advancing research in a broader range of bacteria.