Summary
This project focuses on enhancing the CHyMErA genome editing platform by optimizing Cas12a variants, improving hgRNA scaffolds, refining guide design algorithms, and developing robust CRISPRi and CRISPRa systems for diverse genome editing applications.
What they want
The project involves developing cell lines expressing diverse Cas12a variants and conducting comprehensive functional screening using the Cas9-Cas12a CHyMErA platform with an optimized hgRNA library. This work identified Cas12a variants with enhanced precision and editing efficiency, which are now deployed in high-throughput screens to identify alternative exons influencing cellular fitness and essential genes driving cancer cell proliferation. Systematic screens were also conducted to enhance and diversify the hgRNA scaffold for guide multiplexing and higher-order genetic interaction mapping. In parallel, guide design algorithms are being refined to improve on-target scoring accuracy for enhanced Cas12a nucleases. A comprehensive catalog of potential Cas9 and Cas12a target sites across human and mouse genomes has been compiled and systematically scored for on- and off-target characteristics, with data curated in an online database. A user-friendly search interface is under development for rapid design of individual or large-scale guide RNAs. Additionally, multiple CRISPRi and CRISPRa systems have been established in human cell lines, systematically assessed using high-throughput screening, leading to the identification of a preferred CHyMErA-CRISPRi system and confirmation of Cas12a guide scaffold variants for multiplexed CRISPRi applications.
Deliverables
- Comprehensive catalog of potential Cas9 and Cas12a target sites across human and mouse genomes
- Online database of guide scores and genomic context
- User-friendly, efficient search interface for guide RNA design
Technical requirements
- Cas12a variants
- Cas9-Cas12a CHyMErA platform
- Optimized hgRNA library
- CRISPRi and CRISPRa systems
