FlyBase:CRISPR

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CRISPR

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CRISPR gRNA Design Resources

Resource	Description	Author/Source
CRISPR Optimal Target Finder	Identifies gRNA targets within a provided sequence and additionally searches genome-wide (release 6, current FlyBase release) for potential off-target sites. Includes numerous Drosophila species.	O'Connor-Giles, Wildonger, and Harrison Labs University of Wisconsin-Madison WI, USA
CRISPRscan	Searchable and browsable collection of genome-wide (release 6, current FlyBase release) sgRNA target sites. Displays sgRNA sites as tracks on UCSC's genome browser and allows searching via gene or prediction via sequence. Support for numerous species (fly, zebrafish, mouse, human, others).	Giraldez Lab Yale University New Haven, CT, USA
DRSC Find CRISPR Tool	A tool for discovering gRNA targets located throughout the fly genome (release 5) using gene IDs, symbols, or chromosome locations. Allows for filtering of off-target sites and control of mis-match stringency.	Drosophila RNAi Screening Center (DRSC) Harvard Medical School Boston, MA, USA
E-CRISPR	gRNA genome-wide (release 5) target finder, searchable via gene symbol or sequence. Support for numerous species (fly, zebrafish, mouse, human, others). Offers "relaxed, medium and strict" search options.	Boutros lab, DKFZ Heidelberg, Germany
CHOPCHOP	Search for gRNA targets (release 5). Flexible inputs (gene name, genomic coordinates or DNA sequence) and detection of off-target sites.	Schier & Church Labs Harvard University Cambridge, MA, USA
CRISPR Design	Identification of gRNA sites genome wide (release 5). Offers options for multiple species.	Zhang Lab MIT Boston, MA, USA
CRISPOR	A program that helps design, evaluate, and clone guide sequences for the CRISPR/Cas9 system. Allows for searches across multiple species of Drosophila, including melanogaster (release 5).	Tefor France

CRISPR Stocks

Resource	Description	Author/Source
CRISPR system stocks for site-specific mutagenesis	The Bloomington Stock Center's CRISPR/Cas9 page. Includes various stocks for expression of both Cas9 and tracrRNA.	Bloomington Drosophila Stock Center (BDSC) Indiana University Bloomington, IN, USA
CRISPR Fly Design	Information on stocks generated by the CRISPR Fly Design group at the MRC Laboratory of Molecular Biology.	MRC Laboratory of Molecular Biology Cambridge, UK

CRISPR Vectors

Resource	Description	Author/Source
Addgene CRISPR/Cas9 Plasmids for use in insects	A list of available Cas9 & gRNA expression plasmids for use in insects, including Drosophila.	Addgene Cambridge, MA, USA
CRISPR Fly Design Plasmids	Descriptions and information regarding the plasmids created by the CRISPR Fly Design group (also deposited at Addgene).	MRC Laboratory of Molecular Biology Cambridge, UK

Additional Useful CRISPR Links

Resource	Description	Author/Source
CRISPR Fly Design	An overview and resource collection for CRISPR gene editing in Drosophila.	MRC Laboratory of Molecular Biology Cambridge, UK
flyCRISPR	An overview and resource collection for CRISPR gene editing in Drosophila.	O'Connor-Giles, Wildonger, and Harrison Labs University of Wisconsin-Madison WI, USA
Insect Genomic Technologies Research Coordination Network	Discussion and dissemination of advanced technologies for genome modification of insects through symposia, technical workshops, and training fellowships.	Institute for Bioscience and Biotechnology Research University of Maryland Rockville, MD, USA
FlyRNAi Blog, CRISPR tags	A research blog tracking functional genomics techniques in Drosophila, filtered for keyword: CRISPRs.	Drosophila RNAi Screening Center (DRSC) Harvard Medical School Boston, MA, USA
DRSC Single Cell Cloning Protocol	Single-cell cloning method useful for Drosophila cell CRISPR gene editing	Drosophila RNAi Screening Center (DRSC) Harvard Medical School Boston, MA, USA

Selected recent CRISPR method reviews

• Venken KJ, Sarrion-Perdigones A, Vandeventer PJ, Abel NS, Christiansen AE, Hoffman KL. Genome engineering: Drosophila melanogaster and beyond. Wiley Interdiscip Rev Dev Biol. 2015 Oct 8. doi: 10.1002/wdev.214. [Epub ahead of print] Review. PubMed PMID: 26447401.
• Beumer KJ, Carroll D. Targeted genome engineering techniques in Drosophila. Methods. 2014 Jun 15;68(1):29-37. doi: 10.1016/j.ymeth.2013.12.002. Epub 2014 Jan 8. Review. PubMed PMID: 24412316; PubMed Central PMCID: PMC4048800.
• Xu J, Ren X, Sun J, Wang X, Qiao HH, Xu BW, Liu LP, Ni JQ. A Toolkit of CRISPR-Based Genome Editing Systems in Drosophila. J Genet Genomics. 2015 Apr 20;42(4):141-9. doi: 10.1016/j.jgg.2015.02.007. Epub 2015 Mar 12. Review. PubMed PMID: 25953352.

Selected recent CRISPR method reports for tissue culture cells

• Bassett AR, Tibbit C, Ponting CP, Liu JL. Mutagenesis and homologous recombination in Drosophila cell lines using CRISPR/Cas9. Biol Open. 2014 Jan 15;3(1):42-9. doi: 10.1242/bio.20137120. PMID: 24326186.
• Böttcher R, Hollmann M, Merk K, Nitschko V, Obermaier C, Philippou-Massier J, Wieland I, Gaul U, Förstemann K. Efficient chromosomal gene modification with CRISPR/cas9 and PCR-based homologous recombination donors in cultured Drosophila cells. Nucleic Acids Res. 2014 Jun;42(11):e89. doi: 10.1093/nar/gku289. Epub 2014 Apr 19. PMID: 24748663
• Bassett AR, Kong L, Liu JL. A genome-wide CRISPR library for high-throughput genetic screening in Drosophila cells. J Genet Genomics. 2015 Jun 20;42(6):301-9. doi: 10.1016/j.jgg.2015.03.011. Epub 2015 Apr 18. PMID: 26165496
• Housden BE, Valvezan AJ, Kelley C, Sopko R, Hu Y, Roesel C, Lin S, Buckner M, Tao R, Yilmazel B, Mohr SE, Manning BD, Perrimon N. Identification of potential drug targets for tuberous sclerosis complex by synthetic screens combining CRISPR-based knockouts with RNAi. Sci Signal. 2015 Sep 8;8(393):rs9. doi: 10.1126/scisignal.aab3729. PMID: 26350902

Selected recent CRISPR method reports for in vivo

• Gratz SJ, Cummings AM, Nguyen JN, Hamm DC, Donohue LK, Harrison MM, Wildonger J, O'Connor-Giles KM. Genome engineering of Drosophila with the CRISPR RNA-guided Cas9 nuclease. Genetics. 2013 Aug;194(4):1029-35. doi: 10.1534/genetics.113.152710. Epub 2013 May 24. PMID: 23709638

• Bassett AR, Tibbit C, Ponting CP, Liu JL. Highly efficient targeted mutagenesis of Drosophila with the CRISPR/Cas9 system. Cell Rep. 2013 Jul 11;4(1):220-8. doi: 10.1016/j.celrep.2013.06.020. Epub 2013 Jul 1. PMID: 23827738

• Yu Z, Ren M, Wang Z, Zhang B, Rong YS, Jiao R, Gao G. Highly efficient genome modifications mediated by CRISPR/Cas9 in Drosophila. Genetics. 2013 Sep;195(1):289-91. doi: 10.1534/genetics.113.153825. Epub 2013 Jul 5. PMID: 23833182

• Kondo S, Ueda R. Highly improved gene targeting by germline-specific Cas9 expression in Drosophila. Genetics. 2013 Nov;195(3):715-21. doi: 10.1534/genetics.113.156737. Epub 2013 Sep 3. PMID: 24002648

• Ren X, Sun J, Housden BE, Hu Y, Roesel C, Lin S, Liu LP, Yang Z, Mao D, Sun L, Wu Q, Ji JY, Xi J, Mohr SE, Xu J, Perrimon N, Ni JQ. Optimized gene editing technology for Drosophila melanogaster using germ line-specific Cas9. Proc Natl Acad Sci U S A. 2013 Nov 19;110(47):19012-7. doi: 10.1073/pnas.1318481110. Epub 2013 Nov 4. PMID: 24191015

• Port F, Muschalik N, Bullock SL. Systematic Evaluation of Drosophila CRISPR Tools Reveals Safe and Robust Alternatives to Autonomous Gene Drives in Basic Research. G3 (Bethesda). 2015 May 20;5(7):1493-502. doi: 10.1534/g3.115.019083. PMID: 25999583

• Ren X, Yang Z, Xu J, Sun J, Mao D, Hu Y, Yang SJ, Qiao HH, Wang X, Hu Q, Deng P, Liu LP, Ji JY, Li JB, Ni JQ. Enhanced specificity and efficiency of the CRISPR/Cas9 system with optimized sgRNA parameters in Drosophila. Cell Rep. 2014 Nov 6;9(3):1151-62. doi: 10.1016/j.celrep.2014.09.044. Epub 2014 Oct 23. PMID: 25437567

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