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Loss of Function Genetic Screening with Pooled shRNA or sgRNA Libraries

The basis of Cellecta’s loss-of-function genetic screening technology is the stable suppression or knockout of specific genes on a large-scale using pooled shRNA or sgRNA libraries in mammalian cell systems. Genetic screens with shRNA or sgRNA libraries can be utilized to investigate any aspect of biology that can be recapitulated in a cell culture model. As opposed to expressing and assaying the functional effects of an individual siRNA molecule, the development of complex shRNA and sgRNA libraries allows for simultaneous screening of thousands of different molecules on a target population. In general, genetic screens represent an unbiased approach to identify genes that act in specific cellular pathways.

HT loss-of-function genetic screens have been proven to be an extremely potent and versatile tool to explore the molecular basis of cancer development and progression, and to discover genes essential for viability in cancer cells that can be used as targets for anticancer drug development.

Outline of primary shRNA library screen with pooled Pathway DECIPHER lentiviral library

The screening process introduces a packaged lentiviral library encoding a highly heterogeneous population of barcoded shRNA or sgRNA constructs into a population of identical cells under conditions where each transduced cell will express only a single gene-specific siRNA. Cells exhibiting the desired phenotypic changes are isolated, and the shRNA constructs, presumably inducing the phenotypes, are recovered by PCR and identified by HT sequencing of shRNA-specific bar-codes.

For more details on how RNAi genetic screening with pooled shRNA libraries works, please download the Pooled Lentiviral shRNA Library Screening Reference Manual. Also, please review Cellecta’s RNAi Genetic Screening Services.

For CRISPR screening, you may download the CRISPR sgRNA Library User Manual and see the CRISPR Screening Service web page.