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CRISPR screen to determine host factors

Discover host factors critical for SARS-CoV-2 pathogenesis and identify novel therapeutic targets

CRISPR screens in the field of virology are ideally suited for analyzing virus-host interactions; determining host factors critical for virus entry, replication, and spread; and identifying and prioritizing novel drug targets. Performing a genome-scale CRISPR/Cas9 knockout screen with an sgRNA library can help identify important host factors responsible for disease response.

The Guide-it CRISPR Genome-Wide sgRNA Library System is a pooled lentiviral sgRNA library for performing phenotypic knockout screens in human cell lines. The system is based on the Brunello library (Doench et al. 2016; Doench et al. 2018), and contains over 76,000 guide RNAs (sgRNAs) targeting about 19,000 genes, with four highly active guides per gene, as well as nontargeting sgRNA controls. The amplified, validated library is provided as a ready-to-use lyophilized transfection mix enabling easy production of high-titer lentivirus and setup of genome-wide CRISPR screens (Figure 1).

Following screening, next-generation sequencing (NGS) analysis can be performed to identify candidate genes of interest using the Guide-it CRISPR Genome-Wide sgRNA Library NGS Analysis Kit, which includes the necessary primers and reagents to prepare Illumina-sequencing-ready NGS libraries from cell populations transduced with our genome-wide sgRNA library.

Quality control to ensure sgRNA representation

Our library has been carefully manufactured to ensure proper sgRNA representation. We have measured sgRNA representation at every step in the process: library synthesis, amplification, viral production, and target cell transduction (Figure 2).

Representation of sgRNAs within the system

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632651

Guide-it™ CRISPR Genome-Wide Library PCR Kit

20 Rxns

USD $349.00

The Guide-it CRISPR Genome-Wide Library PCR Kit is designed for NGS library preparation from cell populations screened using the Guide-it CRISPR Genome-Wide sgRNA Library System (Cat. # 632646), and is also sold as part of the Guide-it CRISPR Genome-Wide sgRNA Library NGS Analysis Kit (Cat. # 632647). The kit contains all the primers and reagents necessary for amplification of proviral sgRNA sequences from screened cell populations for analysis by NGS. This kit includes enough reagents for performing 20 PCRs.

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Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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632646

Guide-it™ CRISPR Genome-Wide sgRNA Library System

5 Screens

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63	Use of this product is covered by one or more of the following U.S. Patent Nos. and corresponding patent claims outside the U.S.: 8,562,966, 8,557,231. This product is intended for research purposes only. It may not be used for (i) any human or veterinary use, including without limitation therapeutic and prophylactic use, (ii) any clinical use, including without limitation diagnostic use, (iii) screening of chemical and/or biological compounds for the identification of pharmaceutically active agents (including but not limited to screening of small molecules), target validation, preclinical testing services, or drug development. Any use of this product for any of the above mentioned purposes requires a license from the Massachusetts Institute of Technology.
272	This product (“Product”) and its use, is the subject of U.S. Patents 8,697,359 and 8,771,945 and pending U.S. Patent applications. The purchase of the Product conveys to the buyer the non-transferable right to use Product(s) purchased from Takara Bio USA, Inc. or its Affiliates, and any progeny, modification or derivative of a Product, or any cell or animal made or modified through use of a Product, or any progeny, modification or derivative of such cell or animal (“Related Material”), solely for research conducted by the buyer in accordance with all of the following requirements. No right is given to use this Product or Related Material for any other purpose, including, but not limited to, use in drugs, in vitro diagnostic purposes, therapeutics, or in humans. (i) The buyer shall not sell or otherwise transfer Products (including without limitation any material that contains a Licensed Product in whole or part) or any Related Material to any other person or entity, or use Products or any Related Material to perform services for the benefit of any other person or entity, (ii) the buyer shall use only the purchased amount of the Products and components of the Products, and shall use any Related Material, only for its internal research and not for (a) the practice, performance or provision of any method, process or service, or (b) the manufacture, sale, use, distribution, disposition or importing of any product, in each case (a) or (b) for consideration, or on any other commercial basis (“Commercial Purpose”), (iii) the buyer shall use Licensed Products and any Related Material in compliance with all applicable laws and regulations, including without limitation applicable human health and animal welfare laws and regulations, and (iv) the buyer shall indemnify, defend and hold harmless MIT, Harvard and The Broad and their current and former trustees, directors, officers, faculty, affiliated investigators, students, employees, and agents and their respective successors, heirs and assigns (“Indemnitees”), against any liability, damage, loss, or expense (including without limitation reasonable attorneys’ fees and expenses) incurred by or imposed upon any of the Indemnitees in connection with any claims, suits, investigations, actions, demands or judgments arising out of or related to the exercise of any rights granted to the buyer, or any breach of the rights granted hereunder by the buyer.
406	This product is the subject of a technology license agreement for internal research use only. Use of this product other than for research use may require additional licenses. Information on license restrictions or for uses other than research may be obtained by contacting licensing@takarabio.com.
89	This Product is protected by one or more patents from the family consisting of: US9945850 and any corresponding patents, divisionals, continuations, patent applications and foreign filings sharing common priority with the same family.
330	This product is the subject of a technology license agreement for internal research use only. Use of this product other than for research use may require additional licenses. Information on license restrictions or for uses other than research may be obtained by contacting licensing@takarabio.com.
391	LIMITED USE LABEL LICENSE: RESEARCH USE ONLY Notice to Purchaser: This product is the subject to a license granted to Takara Bio USA, Inc. and its Affiliates from Caribou Biosciences, Inc., and this product is transferred to the end-user purchaser (“Purchaser”) subject to a “Limited Use Label License” conveying to the Purchaser a limited, non-transferable right to use the product, solely as provided to Purchaser, together with (i) progeny or derivatives of the product generated by the Purchaser (including but not limited to cells), and (ii) biological material extracted or derived from the product or its corresponding progeny or derivatives (including but not limited to cells) (collectively, the product, and (i) and (ii) are referred to as (“Material”) only to perform internal research for the sole benefit of the Purchaser. The Purchaser cannot sell or otherwise transfer Material to a third party or otherwise use the Material for any Excluded Use. “Excluded Use” means any and all: (a) commercial activity including, but not limited to, any use in manufacturing (including but not limited to cell line development for purposes of bioproduction), product testing, or quality control; (b) preclinical or clinical testing or other activity directed toward the submission of data to the U.S. Food and Drug Administration, or any other regulatory agency in any country or jurisdiction where the active agent in such studies comprises the Material; (c) use to provide a service, information, or data to a third party; (d) use for human or animal therapeutic, diagnostic, or prophylactic purposes or as a product for therapeutics, diagnostics, or prophylaxis; (e) activity in an agricultural field trial or any activity directed toward the submission of data to the U.S. Department of Agriculture or any other agriculture regulatory agency; (f) high throughput screening drug discovery purposes (i.e., the screening of more than 10,000 experiments per day) as well as scale-up production activities for commercialization; (g) modification of human germline, including editing of human embryo genomes (with the sole exception of editing human embryonic stem (ES) cell lines for research purposes) or reproductive cells; (h) self-editing; and/or (i) stimulation of biased inheritance of a particular gene or trait or set of genes or traits (“gene drive”). It is the Purchaser’s responsibility to use the Material in accordance with all applicable laws and regulations. For information on obtaining additional rights, including commercial rights, please contact licensing@cariboubio.com or Caribou Biosciences, Inc., 2929 7th Street, Suite 105, Berkeley, CA 94710 USA, Attn: Licensing.
396	Sigma-Aldrich CRISPR Use License Agreement This Product and its use are the subject of one or more of the following issued patents and patent applications: Australia Patent Nos. 2013355214; 2017204031; and 2018229489; Canada Patent Nos. 2,891,347 and 2,977,152; China Patent No. CN105142669; European Patent Nos. EP 2 928 496 B1; EP 3 138 910 B1, 3 138 911 B1, EP 3 138 912 B1, EP 3 360 964 B1, EP 3 363 902 B1; Israel Patent No. IL238856; Singapore Patent No. 11201503824S; South Korea Patent Nos. 10-1844123 and 10-2006880; and U.S. Patent Application Serial Nos. 15/188,911; 15/188,924; 15/188,927; 15/188,931; and 15/456,204 (the “Patent Rights”). The purchase of this Product conveys to you (the “Buyer”) the NON-TRANSFERABLE right to use the Product for Licensed Research Use (see definition below) subject to the conditions set out in this License Agreement. 1. “Licensed Research Use” means any use for research purposes, except: (i) Buyer may not sell or otherwise transfer the Product (including without limitation any material that contains the Product in whole or part) or any Related Material to any other third party (except that you may transfer the Product, or any Related Material to a bona fide collaborator or contract research organization), or use the Products or any Related Material to perform services for the benefit of any other third party; (ii) Buyer may use only the purchased amount of the Product and components of the Product, and shall use any Related Material, only for your internal research within the Field, and not for any Commercial Purposes; (iii) Buyer shall use the Product and any Related Material in compliance with all applicable laws and regulations, including without limitation applicable human health and animal welfare laws and regulations; and (iv) the Buyer shall indemnify, defend, and hold harmless SIGMA and their current and former directors, officers, employees and agents, and their respective successors, heirs and assigns (the “Indemnities”) against any liability, damage, loss, or expense (including without limitation reasonable attorneys’ fees and expenses) incurred by or imposed upon any of the Indemnitees in connection with any claims, suits, investigations, actions, demands or judgments arising out of or related to the exercise of any rights granted to the Buyer hereunder or any breach of this License Agreement by such Buyer. 2. For purposes of Section 1 above, the following definitions shall apply: “Commercial Purposes” means (a) the practice, performance or provision of any method, process or service, or (b) the manufacture, sale, use, distribution, disposition or importing of any product, in each case (a) or (b) for consideration, or on any other commercial basis. “Field” means use as a research tool for research purposes; provided, however, that notwithstanding the foregoing, the Field shall expressly exclude (a) any in vivo and ex vivo human or clinical use, including, without limitation, any administration into humans or any diagnostic or prognostic use, (b) the creation of transgenic rodent models and/or derivatives thereof (including, but not limited to, rodents’ cells and rodents’ organs) by for-profit entities, (c) any in vivo veterinary or livestock use, or non-research agricultural use, or (d) use as a testing service, therapeutic or diagnostic for humans or animals. “Related Materials” means any progeny, modification or derivative of a Product. 3. Your right to use the Product will terminate immediately if you fail to comply with these terms and conditions. You shall, upon such termination of your rights, destroy all Product, Related Materials, and components thereof in your control, and notify SIGMA of such in writing. For information on purchasing a license to this Product for purposes other than Licensed Research Use, contact your local SIGMA sales representative, or call +1 800-325-3010.

The Guide-it CRISPR Genome-Wide sgRNA Library System includes all of the components necessary to perform five lentiviral-based, genome-wide CRISPR/Cas9 knockout screens in human cells. Provided with the kit are lentiviral vectors for expression of Cas9 and >76,000 sgRNAs targeting >19,000 human genes (four guides per gene) in lyophilized, ready-to-transfect formats that simplify the production of high-titer lentivirus. The sgRNA library included with this kit is based on the Brunello library (Doench et al. 2016), and incorporates design features intended to maximize on-target efficacy and minimize off-target effects. To ensure balanced guide representation, we analyze the sgRNA library by NGS to confirm that >90% of the included sgRNAs fall within a 10-fold distribution range. Enrichment or depletion of sgRNAs in screened cells can be analyzed by NGS with the Guide-it CRISPR Genome-Wide sgRNA Library NGS Analysis Kit (Cat. # 632647).

Components

632646

Guide-it™ CRISPR Genome-Wide sgRNA Library System

5 Screens

Inquire for Quotation

License Statement

ID Number
63	Use of this product is covered by one or more of the following U.S. Patent Nos. and corresponding patent claims outside the U.S.: 8,562,966, 8,557,231. This product is intended for research purposes only. It may not be used for (i) any human or veterinary use, including without limitation therapeutic and prophylactic use, (ii) any clinical use, including without limitation diagnostic use, (iii) screening of chemical and/or biological compounds for the identification of pharmaceutically active agents (including but not limited to screening of small molecules), target validation, preclinical testing services, or drug development. Any use of this product for any of the above mentioned purposes requires a license from the Massachusetts Institute of Technology.
272	This product (“Product”) and its use, is the subject of U.S. Patents 8,697,359 and 8,771,945 and pending U.S. Patent applications. The purchase of the Product conveys to the buyer the non-transferable right to use Product(s) purchased from Takara Bio USA, Inc. or its Affiliates, and any progeny, modification or derivative of a Product, or any cell or animal made or modified through use of a Product, or any progeny, modification or derivative of such cell or animal (“Related Material”), solely for research conducted by the buyer in accordance with all of the following requirements. No right is given to use this Product or Related Material for any other purpose, including, but not limited to, use in drugs, in vitro diagnostic purposes, therapeutics, or in humans. (i) The buyer shall not sell or otherwise transfer Products (including without limitation any material that contains a Licensed Product in whole or part) or any Related Material to any other person or entity, or use Products or any Related Material to perform services for the benefit of any other person or entity, (ii) the buyer shall use only the purchased amount of the Products and components of the Products, and shall use any Related Material, only for its internal research and not for (a) the practice, performance or provision of any method, process or service, or (b) the manufacture, sale, use, distribution, disposition or importing of any product, in each case (a) or (b) for consideration, or on any other commercial basis (“Commercial Purpose”), (iii) the buyer shall use Licensed Products and any Related Material in compliance with all applicable laws and regulations, including without limitation applicable human health and animal welfare laws and regulations, and (iv) the buyer shall indemnify, defend and hold harmless MIT, Harvard and The Broad and their current and former trustees, directors, officers, faculty, affiliated investigators, students, employees, and agents and their respective successors, heirs and assigns (“Indemnitees”), against any liability, damage, loss, or expense (including without limitation reasonable attorneys’ fees and expenses) incurred by or imposed upon any of the Indemnitees in connection with any claims, suits, investigations, actions, demands or judgments arising out of or related to the exercise of any rights granted to the buyer, or any breach of the rights granted hereunder by the buyer.
406	This product is the subject of a technology license agreement for internal research use only. Use of this product other than for research use may require additional licenses. Information on license restrictions or for uses other than research may be obtained by contacting licensing@takarabio.com.
89	This Product is protected by one or more patents from the family consisting of: US9945850 and any corresponding patents, divisionals, continuations, patent applications and foreign filings sharing common priority with the same family.
330	This product is the subject of a technology license agreement for internal research use only. Use of this product other than for research use may require additional licenses. Information on license restrictions or for uses other than research may be obtained by contacting licensing@takarabio.com.
391	LIMITED USE LABEL LICENSE: RESEARCH USE ONLY Notice to Purchaser: This product is the subject to a license granted to Takara Bio USA, Inc. and its Affiliates from Caribou Biosciences, Inc., and this product is transferred to the end-user purchaser (“Purchaser”) subject to a “Limited Use Label License” conveying to the Purchaser a limited, non-transferable right to use the product, solely as provided to Purchaser, together with (i) progeny or derivatives of the product generated by the Purchaser (including but not limited to cells), and (ii) biological material extracted or derived from the product or its corresponding progeny or derivatives (including but not limited to cells) (collectively, the product, and (i) and (ii) are referred to as (“Material”) only to perform internal research for the sole benefit of the Purchaser. The Purchaser cannot sell or otherwise transfer Material to a third party or otherwise use the Material for any Excluded Use. “Excluded Use” means any and all: (a) commercial activity including, but not limited to, any use in manufacturing (including but not limited to cell line development for purposes of bioproduction), product testing, or quality control; (b) preclinical or clinical testing or other activity directed toward the submission of data to the U.S. Food and Drug Administration, or any other regulatory agency in any country or jurisdiction where the active agent in such studies comprises the Material; (c) use to provide a service, information, or data to a third party; (d) use for human or animal therapeutic, diagnostic, or prophylactic purposes or as a product for therapeutics, diagnostics, or prophylaxis; (e) activity in an agricultural field trial or any activity directed toward the submission of data to the U.S. Department of Agriculture or any other agriculture regulatory agency; (f) high throughput screening drug discovery purposes (i.e., the screening of more than 10,000 experiments per day) as well as scale-up production activities for commercialization; (g) modification of human germline, including editing of human embryo genomes (with the sole exception of editing human embryonic stem (ES) cell lines for research purposes) or reproductive cells; (h) self-editing; and/or (i) stimulation of biased inheritance of a particular gene or trait or set of genes or traits (“gene drive”). It is the Purchaser’s responsibility to use the Material in accordance with all applicable laws and regulations. For information on obtaining additional rights, including commercial rights, please contact licensing@cariboubio.com or Caribou Biosciences, Inc., 2929 7th Street, Suite 105, Berkeley, CA 94710 USA, Attn: Licensing.
396	Sigma-Aldrich CRISPR Use License Agreement This Product and its use are the subject of one or more of the following issued patents and patent applications: Australia Patent Nos. 2013355214; 2017204031; and 2018229489; Canada Patent Nos. 2,891,347 and 2,977,152; China Patent No. CN105142669; European Patent Nos. EP 2 928 496 B1; EP 3 138 910 B1, 3 138 911 B1, EP 3 138 912 B1, EP 3 360 964 B1, EP 3 363 902 B1; Israel Patent No. IL238856; Singapore Patent No. 11201503824S; South Korea Patent Nos. 10-1844123 and 10-2006880; and U.S. Patent Application Serial Nos. 15/188,911; 15/188,924; 15/188,927; 15/188,931; and 15/456,204 (the “Patent Rights”). The purchase of this Product conveys to you (the “Buyer”) the NON-TRANSFERABLE right to use the Product for Licensed Research Use (see definition below) subject to the conditions set out in this License Agreement. 1. “Licensed Research Use” means any use for research purposes, except: (i) Buyer may not sell or otherwise transfer the Product (including without limitation any material that contains the Product in whole or part) or any Related Material to any other third party (except that you may transfer the Product, or any Related Material to a bona fide collaborator or contract research organization), or use the Products or any Related Material to perform services for the benefit of any other third party; (ii) Buyer may use only the purchased amount of the Product and components of the Product, and shall use any Related Material, only for your internal research within the Field, and not for any Commercial Purposes; (iii) Buyer shall use the Product and any Related Material in compliance with all applicable laws and regulations, including without limitation applicable human health and animal welfare laws and regulations; and (iv) the Buyer shall indemnify, defend, and hold harmless SIGMA and their current and former directors, officers, employees and agents, and their respective successors, heirs and assigns (the “Indemnities”) against any liability, damage, loss, or expense (including without limitation reasonable attorneys’ fees and expenses) incurred by or imposed upon any of the Indemnitees in connection with any claims, suits, investigations, actions, demands or judgments arising out of or related to the exercise of any rights granted to the Buyer hereunder or any breach of this License Agreement by such Buyer. 2. For purposes of Section 1 above, the following definitions shall apply: “Commercial Purposes” means (a) the practice, performance or provision of any method, process or service, or (b) the manufacture, sale, use, distribution, disposition or importing of any product, in each case (a) or (b) for consideration, or on any other commercial basis. “Field” means use as a research tool for research purposes; provided, however, that notwithstanding the foregoing, the Field shall expressly exclude (a) any in vivo and ex vivo human or clinical use, including, without limitation, any administration into humans or any diagnostic or prognostic use, (b) the creation of transgenic rodent models and/or derivatives thereof (including, but not limited to, rodents’ cells and rodents’ organs) by for-profit entities, (c) any in vivo veterinary or livestock use, or non-research agricultural use, or (d) use as a testing service, therapeutic or diagnostic for humans or animals. “Related Materials” means any progeny, modification or derivative of a Product. 3. Your right to use the Product will terminate immediately if you fail to comply with these terms and conditions. You shall, upon such termination of your rights, destroy all Product, Related Materials, and components thereof in your control, and notify SIGMA of such in writing. For information on purchasing a license to this Product for purposes other than Licensed Research Use, contact your local SIGMA sales representative, or call +1 800-325-3010.

Notice to purchaser

Components

632647

Guide-it™ CRISPR Genome-Wide sgRNA Library NGS Analysis Kit

10 Rxns

USD $725.00

The Guide-it CRISPR Genome-Wide sgRNA Library NGS Analysis Kit enables preparation of next-generation sequencing (NGS) libraries from cell populations screened using the Guide-it CRISPR Genome-Wide sgRNA Library System (Cat. # 632646), allowing for identification of sgRNA sequences and corresponding gene knockouts that have increased or decreased in frequency under the conditions of the screen. The kit provides all the components necessary to prepare ten sequencing libraries (20 PCRs), including a genomic DNA purification kit, primers, PCR enzyme and buffers, and a PCR cleanup kit.

Notice to purchaser

Components

*You must be logged in to a Purchasing Account in order to purchase these products online, since the purchase of these products may be restricted depending on your account type. Researchers at not-for-profit accounts receive a limited use license with their purchase of the product. Researchers at for-profit accounts must obtain a license prior to purchase. For details please contact licensing@takarabio.com.

More Information

Examples of studies using genome-wide CRISPR screens for virology

Flint, M. A genome-wide CRISPR screen identifies N-acetylglucosamine-1-phosphate transferase as a potential antiviral target for Ebola virus. Nat. Comm. 285, (2019).

Gebre, M. et al. CRISPR-Cas9 Genetic Analysis of Virus-Host Interactions. Viruses 10, 55 (2018).

Han, J. Genome-wide CRISPR/Cas9 Screen Identifies Host Factors Essential for Influenza Virus Replication. Cell Rep. 23, 596–607 (2018).

Li, B. et al. Genome-wide CRISPR screen identifies host dependency factors for influenza A virus infection. Nat. Comm. 11, 164 (2020).

Marceau, C. D. Genetic dissection of Flaviviridae host factors through genome-scale CRISPR screens. Nature 535, 159–163 (2016).

Park, R. J. A genome-wide CRISPR screen identifies a restricted set of HIV host dependency factors. Nat. Genet. 49, 193–203 (2017).

Puschnik, A. S. et al. A CRISPR toolbox to study virus-host interactions. Nat. Rev. Microbiol. 15, 351–364 (2017).

References

Doench, J. G. et al. Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9. Nat. Biotechnol. 34, 184–191 (2016).

Doench, J. G. et al. Am I ready for CRISPR? A user's guide to genetic screens. Nat. Rev. Genet. 19, 67–80 (2018).

Additional product information

Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.

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CRISPR screen to determine host factors

Discover host factors critical for SARS-CoV-2 pathogenesis and identify novel therapeutic targets

Quality control to ensure sgRNA representation

Notice to purchaser

632651: Guide-it CRISPR Genome-Wide Library PCR Kit

Representation of sgRNAs within the Guide-it CRISPR Genome-Wide sgRNA Library System

Identification and analysis of sgRNAs isolated from cells after a 6-thioguanine screen

Workflow schematic for an sgRNA library screen using 6-thioguanine (6-TG) selection

Determination of transduction efficiency of the sgRNA library lentivirus using mCherry fluorescence

Vector and sgRNA scaffold design used in the Guide-it CRISPR sgRNA library

Notice to purchaser

Representation of sgRNAs within the Guide-it CRISPR Genome-Wide sgRNA Library System

Identification and analysis of sgRNAs isolated from cells after a 6-thioguanine screen

Workflow schematic for an sgRNA library screen using 6-thioguanine (6-TG) selection

Determination of transduction efficiency of the sgRNA library lentivirus using mCherry fluorescence

Vector and sgRNA scaffold design used in the Guide-it CRISPR sgRNA library

Notice to purchaser

Representation of sgRNAs within the Guide-it CRISPR Genome-Wide sgRNA Library System

Identification and analysis of sgRNAs isolated from cells after a 6-thioguanine screen

Workflow schematic for an sgRNA library screen using 6-thioguanine (6-TG) selection

Determination of transduction efficiency of the sgRNA library lentivirus using mCherry fluorescence

Vector and sgRNA scaffold design used in the Guide-it CRISPR sgRNA library

632647: Guide-it CRISPR Genome-Wide sgRNA Library NGS Analysis Kit

More Information

Examples of studies using genome-wide CRISPR screens for virology

References

Additional product information

Find more products for COVID-19 research