{{short description|Computer software that aids design of guide RNAs for CRISPR gene editing}} '''CRISPR-Cas design tools''' are computer software platforms and bioinformatics tools used to facilitate the design of guide RNAs (gRNAs) for use with the CRISPR/Cas gene editing system. __TOC__
==CRISPR-Cas== The CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR associated nucleases) system was originally discovered to be an acquired immune response mechanism used by archaea and bacteria. It has since been adopted for use as a tool in the genetic engineering of higher organisms.
Designing an appropriate gRNA is an important element of genome editing with the CRISPR/Cas system. A gRNA can and at times does have unintended interactions ("off-targets") with other locations of the genome of interest. For a given candidate gRNA, these tools report its list of potential off-targets in the genome thereby allowing the designer to evaluate its suitability prior to embarking on any experiments.
Scientists have also begun exploring the mechanics of the CRISPR/Cas system and what governs how good, or active, a gRNA is at directing the Cas nuclease to a specific location of the genome of interest.<ref name="pmid25184501">{{cite journal|display-authors=6|vauthors=Doench JG, Hartenian E, Graham DB, Tothova Z, Hegde M, Smith I, Sullender M, Ebert BL, Xavier RJ, Root DE|date=December 2014|title=Rational design of highly active sgRNAs for CRISPR-Cas9-mediated gene inactivation|journal=Nature Biotechnology|volume=32|issue=12|pages=1262–7|doi=10.1038/nbt.3026|pmc=4262738|pmid=25184501}}</ref><ref name=":0">{{cite journal | vauthors = Chari R, Mali P, Moosburner M, Church GM | title = Unraveling CRISPR-Cas9 genome engineering parameters via a library-on-library approach | journal = Nature Methods | volume = 12 | issue = 9 | pages = 823–6 | date = September 2015 | pmid = 26167643 | pmc = 5292764 | doi = 10.1038/nmeth.3473 }}</ref> As a result of this work, new methods of assessing a gRNA for its 'activity' have been published,<ref name="pmid25184501" /><ref name=":0" /> and it is now best practice to consider both the unintended interactions of a gRNA as well as the predicted activity of a gRNA at the design stage.
==Table==
The below table lists available tools and their attributes.
{| class="wikitable sortable" |+ List of CRISPR-Cas design tools |- ! scope="col" | Tool Name ! scope="col" | Provider ! scope="col" | Searches whole genome for targets ! scope="col" | Returns all targets of genome ! scope="col" | Seed span and location can be defined ! scope="col" | Maximum number of mismatches supported !Predicts gRNA activity ! scope="col" | Available Protospacer adjacent motif (PAM) sequences ! scope="col" | Annotation is reported ! scope="col" | gRNA suggestion or scoring ! scope="col" class="unsortable" | {{Refh}} |- |CRISPRon, CRISPRoff |[https://rth.dk/resources/crispr/ Center for non-coding RNA in Technology and Health, University of Copenhagen] |Yes |Yes |Yes |All |Yes |NGG, NGA, NAG |Yes |Yes |,<ref>{{cite journal | vauthors = Xiang X, Corsi, GI, Anthon C, Qu K, Pan X, Liang X, Han P, Dong Z, Liu L, Zhong J, Ma T, Wang J, Zhang X, Jiang H, Xu F, Liu X, Xu X, Wang J, Yang H, Bolund L, Church GM, Lin L, Gorodkin J, Luo Y | title = Enhancing CRISPR-Cas9 gRNA efficiency prediction by data integration and deep learning | journal = Nature Communications | volume = 12 | issue = 1 | date = May 2021 | page = 3238 | doi = 10.1038/s41467-021-23576-0 | pmid = 34050182 | pmc = 8163799 | bibcode = 2021NatCo..12.3238X | doi-access = free }}</ref><ref>{{cite journal | vauthors = Alkan F, Wenzel A, Anthon C, Havgaard JH, Gorodkin J | title = CRISPR-Cas9 off-targeting assessment with nucleic acid duplex energy parameters | journal = Genome Biology | volume = 19 | issue = 177 | date = October 2018 | page = 177 | pmid = 30367669 | pmc = 6203265 | doi = 10.1186/s13059-018-1534-x | doi-access = free}}</ref> |- |Invitrogen TrueDesign Genome Editor |[https://apps.thermofisher.com/apps/genome-editing-portal/ Thermo Fisher Scientific] |Yes |Yes |No |3 |No |NGG |Yes |Yes |<ref>{{cite journal | vauthors = Liang X, Potter J, Kumar S, Ravinder N, Chesnut JD | title = Enhanced CRISPR/Cas9-mediated precise genome editing by improved design and delivery of gRNA, Cas9 nuclease, and donor DNA | journal = Journal of Biotechnology | volume = 241 | pages = 136–146 | date = January 2017 | pmid = 27845164 | doi = 10.1016/j.jbiotec.2016.11.011 | doi-access = free }}</ref> |- |Breaking-Cas |Spanish National Center for Biotechnology |Yes (over 1000 genomes) |Yes |Yes (by weights) |4 |No |User customizable |Yes |Yes |<ref>{{cite journal | vauthors = Oliveros JC, Franch M, Tabas-Madrid D, San-León D, Montoliu L, Cubas P, Pazos F | title = Breaking-Cas-interactive design of guide RNAs for CRISPR-Cas experiments for ENSEMBL genomes | journal = Nucleic Acids Research | volume = 44 | issue = W1 | pages = W267-71 | date = July 2016 | pmid = 27166368 | pmc = 4987939 | doi = 10.1093/nar/gkw407 }}</ref> |-
| Cas-OFFinder ||Seoul National University|| Yes || Yes || No || 0-10 |No|| NGG, NRG, NNAGAAW, NNNNGMTT|| No || Yes ||<ref name="pmid24463181">{{cite journal | vauthors = Bae S, Park J, Kim JS | title = Cas-OFFinder: a fast and versatile algorithm that searches for potential off-target sites of Cas9 RNA-guided endonucleases | journal = Bioinformatics | volume = 30 | issue = 10 | pages = 1473–5 | date = May 2014 | pmid = 24463181 | pmc = 4016707 | doi = 10.1093/bioinformatics/btu048 }}</ref> |- |CASTING |Caagle |Yes |Yes |No |3 |No |NGG and NAG |No |Yes |<ref>{{cite journal | vauthors = Enkler L, Richer D, Marchand AL, Ferrandon D, Jossinet F | title = Genome engineering in the yeast pathogen Candida glabrata using the CRISPR-Cas9 system | journal = Scientific Reports | volume = 6 | article-number = 35766 | date = October 2016 | pmid = 27767081 | pmc = 5073330 | doi = 10.1038/srep35766 | bibcode = 2016NatSR...635766E }}</ref>
|- |CRISPy ||Technical University of Denmark|| Yes || Yes || No || All || No || NGG || Yes || Yes ||<ref>{{cite journal | vauthors = Pedersen LE, Ronda C, Hansen HG, Kallehauge TB, Betenbaugh MJ, Nielsen AT, Kildegaard HF | title = Accelerating genome editing in CHO cells using CRISPR Cas9 and CRISPy | journal = Biotechnology and Bioengineering | volume = 111 | issue = 8 | pages = 1604–1616 | date = August 2014 | pmid = 24827782 | pmc = 4312910 | doi = 10.1002/bit.25233 }}</ref> |-
|- | CCTop ||University of Heidelberg|| Yes || Yes || Partial || 5 (0-5) |Yes|| NGG, NRG, NNGRRT, NNNNGATT, NNAGAAW, NAAAAC || Yes || Yes ||<ref name="pmid25909470">{{cite journal | vauthors = Stemmer M, Thumberger T, Del Sol Keyer M, Wittbrodt J, Mateo JL | title = CCTop: An Intuitive, Flexible and Reliable CRISPR/Cas9 Target Prediction Tool | journal = PLOS ONE | volume = 10 | issue = 4 | article-number = e0124633 | year = 2015 | pmid = 25909470 | pmc = 4409221 | doi = 10.1371/journal.pone.0124633 | bibcode = 2015PLoSO..1024633S | doi-access = free }}</ref> |- | CHOPCHOP ||Harvard University|| Yes || Yes || Partial || 0, 2 |No|| NGG, NNAGAA, NNNNGANN || No || Yes ||<ref name="pmid24861617">{{cite journal | vauthors = Montague TG, Cruz JM, Gagnon JA, Church GM, Valen E | title = CHOPCHOP: a CRISPR/Cas9 and TALEN web tool for genome editing | journal = Nucleic Acids Research | volume = 42 | issue = Web Server issue | pages = W401-7 | date = July 2014 | pmid = 24861617 | pmc = 4086086 | doi = 10.1093/nar/gku410 }}</ref> |- | CHOPCHOP v2 ||University of Bergen|| Yes || Yes || Yes || 3 (0-3) |Yes|| User customizable || Yes || Yes ||<ref name="pmid27185894">{{cite journal | vauthors = Labun K, Montague TG, Gagnon JA, Thyme SB, Valen E | title = CHOPCHOP v2: a web tool for the next generation of CRISPR genome engineering | journal = Nucleic Acids Research | volume = 44 | issue = W1 | pages = W272–6 | date = July 2016 | pmid = 27185894 | pmc = 4987937 | doi = 10.1093/nar/gkw398 }}</ref> |- | CRISPOR ||University of California, Santa Cruz TEFOR|| Yes (over 200 genomes) || Yes || No || 4 |Yes|| NGG, NGA, NGCG, NNAGAA, NGGNG, NNGRRT, NNNRRT, NNNNGMTT, NNNNACA, TTTN || Yes || Yes ||<ref name="pmid27380939">{{cite journal | vauthors = Haeussler M, Schönig K, Eckert H, Eschstruth A, Mianné J, Renaud JB, Schneider-Maunoury S, Shkumatava A, Teboul L, Kent J, Joly JS, Concordet JP | display-authors = 6 | title = Evaluation of off-target and on-target scoring algorithms and integration into the guide RNA selection tool CRISPOR | journal = Genome Biology | volume = 17 | issue = 1 | page = 148 | date = July 2016 | pmid = 27380939 | pmc = 4934014 | doi = 10.1186/s13059-016-1012-2 | doi-access = free }}</ref> |-
| CRISPR Design ||Zhang Lab, MIT|| Yes || No || No || 4 |No|| NGG and NAG || mRNA exons || Yes ||<ref name="pmid23873081">{{cite journal | vauthors = Hsu PD, Scott DA, Weinstein JA, Ran FA, Konermann S, Agarwala V, Li Y, Fine EJ, Wu X, Shalem O, Cradick TJ, Marraffini LA, Bao G, Zhang F | display-authors = 6 | title = DNA targeting specificity of RNA-guided Cas9 nucleases | journal = Nature Biotechnology | volume = 31 | issue = 9 | pages = 827–32 | date = September 2013 | pmid = 23873081 | pmc = 3969858 | doi = 10.1038/nbt.2647 | hdl = 1721.1/102691 }}</ref> |- | CRISPRdirect ||Database Center for Life Science (DBCLS) || Yes (over 200 species) || Yes || No || Any number |No|| NNN || Yes || Yes ||<ref name="pmid25414360">{{cite journal | vauthors = Naito Y, Hino K, Bono H, Ui-Tei K | title = CRISPRdirect: software for designing CRISPR/Cas guide RNA with reduced off-target sites | journal = Bioinformatics | volume = 31 | issue = 7 | pages = 1120–3 | date = April 2015 | pmid = 25414360 | pmc = 4382898 | doi = 10.1093/bioinformatics/btu743 }}</ref> |- | CRISPRscan || Giraldez Lab, Yale|| Yes || Yes || No || 4 |Yes|| NGG, TTTV, TTTN || Yes || Yes ||<ref name="pmid26322839">{{cite journal | vauthors = Moreno-Mateos MA, Vejnar CE, Beaudoin JD, Fernandez JP, Mis EK, Khokha MK, Giraldez AJ | title = CRISPRscan: designing highly efficient sgRNAs for CRISPR-Cas9 targeting in vivo | journal = Nature Methods | volume = 12 | issue = 10 | pages = 982–8 | date = October 2015 | pmid = 26322839 | pmc = 4589495 | doi = 10.1038/nmeth.3543 }}</ref> |- | CRISPRseek ||Bioconductor|| Yes || Yes || No || Any number |No|| User customizable || mRNA exons || Yes ||<ref name="pmid25247697">{{cite journal | vauthors = Zhu LJ, Holmes BR, Aronin N, Brodsky MH | title = CRISPRseek: a bioconductor package to identify target-specific guide RNAs for CRISPR-Cas9 genome-editing systems | journal = PLOS ONE | volume = 9 | issue = 9 | article-number = e108424 | year = 2014 | pmid = 25247697 | pmc = 4172692 | doi = 10.1371/journal.pone.0108424 | bibcode = 2014PLoSO...9j8424Z | doi-access = free }}</ref> |- | DESKGEN ||Desktop Genetics|| Yes || Yes || Yes || Any number |Yes|| Fully user customizable || Yes || Yes ||<ref>{{cite news | title = Desktop Genetics Announces the Launch of DeskGen Gene Editing Platform | journal = American Laboratory | year = 2015 | url = http://www.americanlaboratory.com/173625-Desktop-Genetics-Announces-the-Launch-of-DeskGen-Gene-Editing-Platform/ }}</ref> |- | GuideScan ||GuideScan|| Yes || Yes || Yes || 3 on website and customizable with command line |Yes|| NGG/NAG on website and customizable with command line || Yes || Yes ||<ref name="pmid28263296">{{cite journal | vauthors = Perez AR, Pritykin Y, Vidigal JA, Chhangawala S, Zamparo L, Leslie CS, Ventura A | title = GuideScan software for improved single and paired CRISPR guide RNA design | journal = Nature Biotechnology | volume = 35 | issue = 4 | pages = 347–349 | date = April 2017 | pmid = 28263296 | pmc = 5607865 | doi = 10.1038/nbt.3804 }}</ref> |- |GT-Scan |CSIRO & {{nowrap|EMBL-ABR}} |Yes |Yes |Yes |3 (0-3) |No |User customizable |Links to Ensembl genome browser |Yes |<ref>{{cite journal | vauthors = O'Brien A, Bailey TL | title = GT-Scan: identifying unique genomic targets | journal = Bioinformatics | volume = 30 | issue = 18 | pages = 2673–5 | date = September 2014 | pmid = 24860161 | pmc = 4155256 | doi = 10.1093/bioinformatics/btu354 }}</ref> |- | Off-Spotter ||Thomas Jefferson University|| Yes || Yes || Yes || 0-5 | || NGG, NAG, NNNNACA, NNGRRT (R is A or G) ||mRNA exons, unspliced mRNA, mRNA, 5'UTR, CDS, 3'UTR, unspliced lincRNA, lincRNA || User customizable ||<ref name="pmid25630343">{{cite journal | vauthors = Pliatsika V, Rigoutsos I | title = "Off-Spotter": very fast and exhaustive enumeration of genomic lookalikes for designing CRISPR/Cas guide RNAs | journal = Biology Direct | volume = 10 | issue = 1 | page = 4 | date = January 2015 | pmid = 25630343 | pmc = 4326336 | doi = 10.1186/s13062-015-0035-z | doi-access = free }}</ref> |- | sgRNA Designer ||Broad Institute|| No || No || No || 0 |Yes|| NGG || CDS (if searching by transcript ID) || Yes ||<ref name="pmid25184501" /> |- |Synthego Design Tool |Synthego |Yes (over 120,000 genomes) |No (Optimized for Knockout) |Yes |3 |Yes |NGG |Yes (RefSeq, Ensembl, Gencode) |Yes |<ref>TechCrunch. [https://techcrunch.com/2017/05/16/synthegos-genetic-toolkit-aims-to-make-crispr-more-accessible/ "Synthego's genetic toolkit aims to make CRISPR more accessible | May 2017"] Retrieved 23 January 2018.</ref> |- |TUSCAN ||CSIRO|| No || No || No || 0 || Yes || NGG ||No || Yes ||<ref name="pmid31021206">{{cite journal | vauthors = Wilson LO, Reti D, O'Brien AR, Dunne RA, Bauer DC | title = High Activity Target-Site Identification Using Phenotypic Independent CRISPR-Cas9 Core Functionality | journal = The CRISPR Journal | volume = 1 | issue = 2 | pages = 182–190 | date = April 2018 | pmid = 31021206 | doi = 10.1089/crispr.2017.0021 }}</ref> |- |VARSCOT ||CSIRO|| Yes || Yes || No || 0-8 || Yes || User customizable || No || Yes ||<ref name="pmid31248401">{{cite journal | vauthors = Wilson LO, Hetzel S, Pockrandt C, Reinert K, Bauer DC | title = VARSCOT: variant-aware detection and scoring enables sensitive and personalized off-target detection for CRISPR-Cas9 | journal = BMC Biotechnology | volume = 19 | issue = 1 | page = 40 | date = June 2019 | pmid = 31248401 | pmc = 6598273 | doi = 10.1186/s12896-019-0535-5 | doi-access = free }}</ref> |- |CRISPR Targeted Gene Designer |[https://horizondiscovery.com/products/tools/CRISPR-Targeted-Gene-Designer Horizon Discovery]{{Dead link|date=February 2022 |bot=InternetArchiveBot |fix-attempted=yes }} |Yes, Multiple |yes |yes |4 |Yes |NGG, NNGRRT, YTTV, other |Yes |Yes |(21) |- |GuideMaker |[https://ars.usda.gov United States Department of Agriculture, Agricultural Research Service] |Yes, any user supplied genome |Yes |Yes |0-5 |Yes |Any PAM site and PAM orientation |Yes |Yes |<ref>{{cite journal | vauthors = Poudel R, Rodriguez LT, Reisch CR, Rivers AR | title = GuideMaker: Software to design CRISPR-Cas guide RNA pools in non-model genomes | journal = GigaScience | volume = 11 | date = April 2022 | pmid = 35365834 | doi = 10.1093/gigascience/giac007 | pmc = 8975720 | s2cid = 235700000 | doi-access = free }}</ref> |- |WGE |Wellcome Sanger Institute |Yes, human and mouse reference genomes |Yes |Yes |4 (0-4) |No |NGG |Yes |Yes |<ref>{{cite journal | vauthors = Hodgkins A, Farne A, Perera S, Grego T, Parry-Smith DJ, Skarnes WC, Iyer V | title = WGE: a CRISPR database for genome engineering | journal = Bioinformatics | volume = 31 | issue = 18 | pages = 3078–3080 | date = September 2015 | pmid = 25979474 | doi = 10.1093/bioinformatics/btv308 | doi-access = free | pmc = 4565030 }}</ref> |}
== References == {{Reflist}}
{{DEFAULTSORT:CRISPR Cas Tools}} Category:Genetic engineering Category:Genome editing