CRISP-Seq: Integrating the Power of Single-Cell RNA Sequencing and CRISPR-Pooled Screens (No. T4-1848)
Lead Researcher: Prof. Ido Amit
Single-cell RNA sequencing (RNA-Seq) is a powerful tool to identify and characterize the transcriptome of single cells. While providing unprecedented resolution in terms of studying cells, single-cell RNA-Seq is functionally a descriptive tool, unless combined with gene manipulation. CRISPR-Cas9 is a genome-editing technology that enables mutating specific and known targets in the genome by using guide RNAs that match the desired target site. CRISPR can therefore be used to generate single gene knock-outs and to create pooled screens that connect genes to functions. However, single-cell RNA-Seq is not scalable, and CRISPR lacks the resolution to elucidate complex phenotypes.
The research team of Prof. Amit developed a new method which combines the two aforementioned techniques – CRISP-Seq. The method uses the ability of CRISPR-Cas9 to induce site-specific mutations and the power of single-cell RNA-Seq to study gene expression in high resolution. Together they enable to examine gene circuits, pathways and functions affected by interference with numerous genes in a single experiment.
* Enables both manipulation and study of cells
* High resolution of single-cell gene alterations
* High-throughput data on multiple genes from one experiment
CRISP-Seq protocol functions by inducing mutations for one or more target genes, on the single cell level using the CRISPR technology. The vectors used for mutation are barcoded for detection and tracking by RNA-Seq, as well as include probes (e.g. fluorescence) for a visible phenotype that can be used for sorting in flow cytometry. Single-cell RNA-Seq completes the process where the effects on the transcriptome of single or multiple mutations can be examined in high-throughput single cell level.