A whole new way to do functional genomics screens!
CRISPR-Cas9 lentiviral sgRNA pooled screening changes EVERYTHING
Options for functional genomics screening
For over a decade, researchers who want to perform functional genomic screens have had two reagent choices: siRNA and shRNA. For screening, siRNA must be used in an arrayed format – one reagent per well, and this requires robotic liquid handling and high-throughput cell analysis instruments. For this reason arrayed siRNA screening is not an option for investigators who do not have access to this type of equipment.
shRNA screens on the other hand can be conducted in either an arrayed format or a pooled format. Pooled screens differ from arrayed screens in that all of the reagents (shRNAs) are combined into one tube. Analysis of pooled screens also differs because all cells are combined rather than in individual wells as for arrayed screens. A pooled screen of thousands of genes can simply be conducted in a few cell culture dishes as opposed to an arrayed screen which would require many thousands of wells (multiple shRNAs per gene), robotic liquid handling and high-throughput screening. Pooled screening works by comparing the representation of shRNA constructs in control samples to experimentally treated samples. In this way, genes involved in the observed phenotype are identified. Usually, this involves next-generation sequencing.
Welcome, CRISPR-Cas9 lentiviral sgRNA pooled screening, this changes EVERYTHING!
Much has been written about how CRISPR-Cas9 technology is not an evolutionary step but a revolutionary step in molecular biology. A similar argument can be made that CRISPR-Cas9 changes functional genomics screening in ways we could not have anticipated just a short time ago. One key difference is that with CRISPR-Cas9 systems, the potential exists for total ablation of target gene expression as opposed to the partial knockdown observed in most RNAi approaches. This fact alone changes everything from experimental design to analysis and interpretation of results.
Now you can perform phenotypic gene knockout screens
With Edit-R™ Lentiviral sgRNA Pooled Screening Libraries you can perform powerful loss-of-function screens in human, mouse or rat cells. Pre-designed sgRNA libraries target gene families (e.g. kinases, phosphatases, ion channels, etc.), druggable targets or whole genome scales, and custom libraries can also be created for your specific gene lists.
All Edit-R pooled lentiviral sgRNA libraries feature:
- Increased functional knockout
sgRNAs designed by a functionally validated algorithm
- Unparalleled sgRNA specificity
through proprietary, optimized alignment bioinformatics
- Rigorous production standards
strict quality-controlled conditions for low variation in sgRNA representation
- High sensitivity screens
concentrated lentiviral particles (minimum titer of = 5 x 108 TU/mL) and enough volume for high average sgRNA fold representation
- High hit confidence
deep and broad coverage of 5 to 10 sgRNAs per gene
- Inter- and intra-pool normalization
over 400 positive and negative sgRNA controls in each pool
To help you design your CRISPR-Cas9 screening experiments, you are supported by a highly detailed product manual with carefully validated protocols.
For next-generation sequencing analysis, Edit-R Pooled sgRNA Forward and Reverse Index PCR primers have built-in adaptor and index sequences that allow the researcher to easily move from PCR amplification to high-throughput sequencing on the Illumina™ platform. Further, we provide you with a detailed guide for analyzing high-throughput sequencing data and calling hits from pooled sgRNA library screens. Lastly, our experienced team of Technical Support Scientists is always ready to help you.
- Optimized tools for high-confidence genome engineering
- CRISPR-Cas9 systems can be used with custom RNA guides for several gene editing applications