Weekly CRISPR Content

CRISPR Application Highlights – CRISPR Genetic Screens for Target Identification

CRISPR screening is a powerful technique used to identify important genes within a vast number of genetic sequences, such as the entire genome. It can pinpoint genes that influence various physiological effects, including drug resistance and sensitivity. CRISPR-Cas9 is particularly useful for target-specific gene modifications, allowing researchers to identify drug targets or validate and optimize existing ones. By combining CRISPR with next generation sequencing and single-cell sequencing technologies, scientists can perform whole-genome analysis and make significant advancements in identifying disease-specific targets.

The basic principle behind CRISPR screening involves knocking out individual genes in a mixed population of cells to determine their impact on cell survival and growth. CRISPR screening begins with the knockout of one gene per cell via using a CRISPR library contains a pool of lentivirus each possess gene encoding a different gRNA, resulting in a population of cells with different genes knocked out. These cells are then pooled together and allowed to grow for a few days for NGS to identify which genetic sequences are present or knocked out. CRISPR screening libraries are commercially available and enable scientists to analyze specific gene families or pathways by creating targeted gene knockouts and studying the resulting phenotypic alterations.

In certain cases, researchers may have a short list of genes of interest and use an arrayed gRNA CRISPR library. This library consists of synthetic sgRNA kept in separate wells on a multi-well plate. Each well contains one or several predetermined CRISPR targeting sequences that are transfected into cells. This approach eliminates the need for next-generation sequencing but requires automation equipment and is more suitable for screens focused on phenotype changes caused by gene knockout.

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