MIC-Drop: A platform for large-scale in vivo CRISPR screens

Area of Science:

• Genetics
• Developmental Biology
• Bioengineering

Background:

• CRISPR-Cas9 screens are powerful in cell cultures but challenging in animal models due to scalability issues.
• Generating and tracking numerous mutant animals for large-scale genetic screens is currently prohibitive.
• Efficient, scalable methods are needed for functional genetic screens in whole organisms.

Purpose of the Study:

• To develop a scalable platform for large-scale CRISPR screens in zebrafish.
• To enable the identification of genes associated with morphological and behavioral phenotypes.
• To facilitate the discovery of novel genes involved in biological processes and drug target identification.

Main Methods:

• Developed Multiplexed Intermixed CRISPR Droplets (MIC-Drop) platform.
• Combined droplet microfluidics, en masse CRISPR ribonucleoprotein injection, and DNA barcoding.
• Applied MIC-Drop for functional genetic screens in zebrafish.

Main Results:

• MIC-Drop successfully enabled large-scale CRISPR screens in zebrafish.
• The platform efficiently identified genes responsible for specific morphological and behavioral phenotypes.
• Demonstrated MIC-Drop's utility in identifying small-molecule targets and discovering novel genes in cardiac development.

Conclusions:

• MIC-Drop is a scalable platform for genome-scale reverse genetic screens in model organisms like zebrafish.
• This technology overcomes previous limitations in animal-based CRISPR screening.
• MIC-Drop opens new avenues for understanding gene function and identifying therapeutic targets.