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Lentiviral and synthetic reagents for targeted gene knockout
Guide RNAs program Cas9 nucleases to cut at a specific genomic location. The design of an effective, functional guide RNA is critical to achieving specific gene knockout. Take advantage of our algorithm for functional, specific guide RNAs and search our predesigned products for your gene of interest.
Pre-designed or custom synthesized for rapid knockout studies across many genes
Pre-designed or custom sgRNA as glycerol stocks and high-titer purified
lentiviral particles for small- and large-scale studies.
All Edit-R predesigned guide RNA (synthetic crRNA and lentiviral sgRNA) are designed with a validated algorithm to improve the likelihood of functional knockout, not just double-strand breaks (DSB). By assessing functional phenotypes for thousands of designs, then validating our design rules in other assay systems, we have established rules for determining target sites that are more likely to give efficient cleavage and functional knockout with high specificity. See the Supporting Data tab for more information.
In addition to expression of the Cas9 nuclease, the CRISPR-Cas9 system requires a specific RNA moiety to recruit and direct the nuclease activity. These guide RNAs take one of two forms:
Figure 1a. Illustration of Cas9 nuclease programmed by the crRNA:tracr complex cutting both strands of genomic DNA 5' of the PAM
Figure 1b. Illustration of Cas9 nuclease programmed by the sgRNA complex cutting both strands of genomic DNA 5' of the PAM
Algorithm-optimized crRNA for genome-wide coverage of human, mouse, or rat genes. Modifications for nuclease resistance improve DNA-free editing. Simply search for your gene!
Edit-R trans-activating CRISPR RNA (tracrRNA) is synthetic, HPLC-purified, long RNA required for use with Edit-R crRNA to form the complex that programs Cas9 nuclease. It is modified for nuclease resistance and can be used with modified or unmodified Edit-R crRNA.
Species-specific crRNAs targeting well-characterized genes, as well as mismatch detection assay primers, to determine the effectiveness of your gene editing conditions for maximal efficiency.
Non-targeting controls to evaluate cellular responses to CRISPR-Cas9 components in the absence of gene target-specific crRNA.
Plated pre-defined collections of popular gene families for arrayed knockout screening
Have a favorite gene list? Customize and order plates of predesigned crRNA for knockout studies in your targets of interest
Algorithm-optimized sgRNA for genome-wide coverage of human, mouse, or rat genes. Provided as high-titer lentiviral particles and glycerol stocks.
Species-specific sgRNAs targeting well-characterized genes to determine the effectiveness of your gene editing conditions for maximal efficiency.
Non-targeting controls to evaluate cellular responses to CRISPR-Cas9 components in the absence of gene target-specific sgRNA
High-titer pooled screening libraries for pre-defined gene sets in human and mouse.
Arrayed collections of lentiviral sgRNA libraries for high-throughput knockout screening across entire human gene families.
Place a custom gRNA order, or design and order your own crRNA or lentiviral sgRNA with our easy-to-use interface.
Design and order a single-strand DNA donor (≤ 150 nt) for insertion, deletion, or other alteration.
Design and order a plasmid DNA donor kit for insertion of an mKate2 fluorescent marker
Rapidly and easily assemble a plasmid donor for HDR
Quickly and efficiently build a HDR donor plasmid
PCR components for Edit-R Plasmid Donor Kits
While the best guide RNA for your experiment may heavily depend on your particular application or cell type, a few basic questions may help to point you in the right direction for product selection.
All Edit-R predesigned guide RNA (synthetic crRNA and lentiviral sgRNA) are designed with a validated algorithm to improve the likelihood of functional knockout, not just double-strand breaks (DSB).
Assessment of functionality of crRNA along the length of the coding sequence of VCP gene indicates no particular pattern as it relates to exonic location, reinforcing the importance of a design algorithm to characterize functional characteristics. Recombinant U2OS Ubi[G76V]-EGFP -Cas9 cells were transfected with 266 different synthetic crRNA:tracrRNA complexes targeting VCP gene along the length of the coding regions of the gene (using DharmaFECT 4 Transfection Reagent at 0.07 µg/well and 50 nM synthetic crRNA:tracrRNA final concentration). After 72 hours the EGFP fluorescence was measured using an Envision plate reader (Perkin Elmer).
10 crRNAs with high functional scores for 10 genes (blue bars) and 10 crRNAs with low functional scores for the same genes (yellow bars) were tested for editing by Next Generation Sequencing. 93% of the high-scoring crRNAs and 32 % of the low scoring crRNAs showed > 40% of editing (indel formation). The Cas9-HEK293T cell line was transfected with 50 nM crRNA:tracrRNA, using 0.25 µL/well of DharmaFECT 1. Seventy-two hours post-transfection, cells were lysed and Nextera transposon-adapted amplicons spanning each crRNA site were generated for every treated sample as well as for a matched control amplicon from untransfected samples. Samples were indexed using the Nextera 96-well index kit and pooled for sequencing on a MiSeq instrument (paired end reads, 2 x 300 length). Reads that passed NGS quality filtering criteria were aligned to the reference file (Bowtie2 v2.1.0). Percent perfect reads were calculated and normalized to the control untransfected samples (Samtools v0.1.12a); the data is presented as normalized percent edited.
U2OS-Proteasome cells with integrated Cas9 (under CAG promoter) were plated in 96-well plates at 10,000 cells per well. 24h after plating, cells were transfected with 25 nM crRNA:tracrRNA using 0.2 µg/well of DF4. Cell were analyzed for apoptosis 48 h after transfection using the ApoONE homogeneous assay (Promega). Shown is a box plot representation of the functionality of crRNAs targeting BCL2L1, PLK1 or WEE1 in ApoONE assay. To generate the box plot, the crRNAs were divided into bottom half (H1) and top half (H2) based on their functional score. The medians, distribution of data between the lower and upper quartile and the minimum and maximum values demonstrate that high-scoring crRNAs have increased functionality.