Human CRISPR Knockout Pooled Library (GeCKO v2)
(Pooled Libraries #1000000048, #1000000049)
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Purpose
The human GeCKO (Genome-Scale CRISPR Knockout) lentiviral pooled libraries target early consecutive exons for genome editing.
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Vector Backbone
- lentiCRISPR v2 (Plasmid #52961) backbone (one-vector system) - expresses Cas9
- lentiGuide-Puro (Plasmid #52963) backbone (two-vector system) - does not express Cas9
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Depositing Labs
Ordering
Item | Catalog # | Description | Quantity | Price (USD) | |||
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Pooled Library | 1000000048 | Human CRISPR KO Library (GeCKO v2) in lentiCRISPR v2 | 1 | $640 | Add to Cart | ||
Pooled Library | 1000000049 | Human CRISPR KO Library (GeCKO v2) in lentiGuide-Puro + lentiCas9-Blast plasmid | 1 | $640 | Add to Cart |
Library Details
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SpeciesHuman
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Genes targeted19,050
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gRNAs123,411
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Controls1,000 per half-library
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Lentiviral Generation3rd
Library Shipping
Each library is delivered in a microcentrifuge tube on blue ice. The tube's contents will not necessarily be frozen. For best results, minimize freeze/thaws.
This library will be delivered as two pooled DNA half-libraries in microcentrifuge tubes labeled A and B.
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Volume∼20 µL
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Concentration50 ng/µL
For the two-vector system ONLY: You will also receive a bacterial stab of the lentiCas9-Blast plasmid (Addgene #52962). Please do NOT order this plasmid in addition to this library.
Resource Information
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Protocols
- For the most up-to-date and detailed instructions on using this library, please see the protocol paper from Joung, et al.
- Library Amplification Protocol (PDF, 276 KB)
- gRNA Cloning in lentiCRISPR v1, lentiCRISPR v2, and lentiGuide-Puro Vectors (PDF, 2.4 MB)
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Depositor Data
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Terms and Licenses
Academic/Nonprofit TermsIndustry Terms- Not Available to Industry
Trademarks- Zeocin® is an InvivoGen trademark.
Depositor Comments
In the library amplification protocol, we describe how to amplify GeCKO v2.0 DNA plasmids to have sufficient quantity to produce lentivirus, while maintaining full library representation. The GeCKO v2 libraries consist of over 100,000 unique gRNAs for gene knock-out in either the human or mouse genome.
Each species-specific library is delivered as two half-libraries (A and B). When used together, the A and B libraries contain 6 sgRNAs per gene (3 sgRNAs in each library). We recommend screening the entire library (A and B) when possible but if adequate representation cannot be obtained with the entire library, screening can be performed with one half-library. Since more cells are needed in the screen as number of constructs in the library increases, this design has the flexibility of screening with just the A library (3 sgRNAs per gene) at a smaller scale or screening the full library (6 sgRNAs per gene).
Both A and B libraries contain 1,000 control sgRNAs designed not to target in the genome. The A library also targets miRNAs (4 sgRNAs per miRNA). Complete sequences and targets of all sgRNAs in the A and B libraries are available in CSV format above.
The two-vector format with the library in lentiGuide-Puro has the advantage of higher titer for the library virus but requires cells to already contain Cas9 (usually genomically integrated using lentiCas9-Blast). There is also a protocol below for cloning custom gRNA sequences of your own design into any of the vectors (lentiCRISPR v1/2 and lentiGuide-Puro).
For additional information, visit Neville Sanjana Lab's Library screening resources website:
http://sanjanalab.org/lib.html
These pooled libraries were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.
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For your Materials & Methods section:
Human GeCKOv2 CRISPR knockout pooled library was a gift from Feng Zhang (Addgene #1000000048)
Human GeCKOv2 CRISPR knockout pooled library was a gift from Feng Zhang (Addgene #1000000049) -
For your References section:
Improved vectors and genome-wide libraries for CRISPR screening. Sanjana NE, Shalem O, Zhang F. Nat Methods. 2014 Aug;11(8):783-4. doi: 10.1038/nmeth.3047. PubMed 25075903