shRNA Core Facility
The Einstein shRNA Core Facility provides a comprehensive set of tools and expertise for modulating gene function using RNA interference, CRISPR/Cas9, and overexpression approaches. The Facility is equipped for cell line studies requiring knockout, knockdown, overexpression, and other types of modulation of gene expression, at scales ranging from individual genes to genome-scale. Facility staff, headed by John Reidhaar-Olson, Ph.D., have extensive academic and industrial experience performing functional genomics screens, and are available for consultation with investigators regarding experimental design and data analysis. Specific services include:
shRNA services: The Facility houses four genome-wide lentiviral shRNA libraries, between them comprising multiple shRNA sequences for most human and mouse genes. Clones from the libraries can be supplied as glycerol stocks or plasmids, or prepared as viral supernatant or concentrated viral stocks by Facility staff. In addition, custom arrayed shRNA screens can be performed with shRNAs targeting particular gene families, pathways, or other gene sets. Pooled shRNA libraries are also available, for unbiased screening of large gene sets.
CRISPR/Cas9 services: The Facility provides several CRISPR/Cas9-based services, permitting a variety of gene modulations. All use lentiviral vectors and inducible Cas9 constructs. Services include sgRNA design and cloning, packaging of sgRNA and Cas9 vectors into lentivirus, and construction of cell lines stably expressing sgRNAs and Cas9. Available approaches include gene knockout, knock-in, transcriptional activation, and transcriptional repression. These CRISPR-based services can be applied to any mammalian species. Pooled CRISPR screens are also available for human and mouse.
Overexpression services: The Facility has two lentiviral ORF libraries, together representing approximately 12,000 human genes, and the genome-wide human and mouse Mammalian Genome Collections.
Screening support services: The Facility provides support for screening with an EnVision Multi-Label plate reader and an Operetta high-content instrument. The EnVision provides well-based measurement of fluorescence intensity, fluoresccence polarization, luminescence, absorbance, and time-resolved fluorescence. The Operetta provides cell-based imaging analysis, and is suitable for RNAi-based and other arrayed screens. The Operetta includes fluorescence and brightfield options, confocal imaging, eight excitation and four emission filters, an environmental control chamber for live-cell imaging, and data analysis software for indentifying and quantitating a wide variety of cellular parameters. Facility staff are available to perform image analysis.