In 2016 the two initial Perturb-seq papers were published in Cell. Single-cell RNA sequencing is used to measure the effects of many CRISPR-based perturbations on large numbers of cells. To identify specific DNA changes or perturbations on individual cells within pooled experiments, cell barcoding is used on single cells from which transcriptional profiles were also acquired. Using droplet-based single-cell RNA sequencing, each cell has a gene expression profile associated one or more genetic perturbations.
Researchers of the paper led by the Broad Institute used CRISPR-Cas9 nucleases to cut DNA and inactivate genes for transcription factors (TFs), proteins that control the expression of genes involved in immune response in dendritic cells. They also inactivated genes for TFs and cell cycle regulator proteins in a cancer cell line.Lead author Jonathan S. Weissman is founder of KSQ therapeutics a company that uses CRISPR for genome screening.
The UCSF-led experiments used CRISPR-based transcriptional interference (CRISPRi) to repress genes in a cancer cell line to investigate errors in protein production that result from the cell sensing stress. Lead author Aviv Regev is a scientific advisor to ThermoFisher, Syros, and Driver.
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- Broad InstituteA biomedical and genomic research center
- CRISPRClustered regularly interspaced short palindromic repeats (CRISPR) is a prokaryotic adaptive immune response that provides immunity against foreign nucleic acids, such as viral DNA and bacterial plasmids, through the use of crRNAs (CRISPR RNAs) and associated Cas genes.
- CRISPR-Cas9CRISPR-Cas9 is a genome editing system. CRISPR systems provides immunity to bacteria and archaea from viruses and has been adapted for use as a genome editing tool capable of knocking out genes and rewriting genetic sequences in animal, plant and fungi. CRISPR-Cas9 is being adapted to other applications outside genome editing.
- CRISPR interferenceA CRISPR technology which uses dCas9 (a Cas9 complex with no endonuclease funtion) and a guide RNA sequence to target and repress function of a DNA region.