CRISPR/Cas Tools
This page describes a list of software platforms and bioinformatics tools built to facilitate the design of guide RNAs (gRNAs) for use with the CRISPR/Cas system.
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.[1][2] As a result of this work, new methods of assessing a gRNA for its 'activity' have been published,[1][2] 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.
The below table lists available tools and their attributes, and includes links to the corresponding websites.
Tool Name | Provider | Searches whole genome for targets | Returns all targets of genome | Seed span and location can be defined | Maximum number of mismatches supported | Predicts gRNA activity | Available Protospacer adjacent motif (PAM) sequences | Annotation is reported | gRNA suggestion or scoring | External Link | References |
---|---|---|---|---|---|---|---|---|---|---|---|
Benchling CRISPR gRNA Design | Benchling | Yes | Yes | Yes | 4 | Yes | User customizable | Yes | Yes | Webserver | - |
Breaking-Cas | Spanish National Center for Biotechnology | Yes | Yes | Yes (by weights) | 4 | No | User customizable | Yes | Yes | Webserver | [3] |
Cas-OFFinder | Seoul National University | Yes | Yes | No | 0-10 | No | NGG, NRG, NNAGAAW, NNNNGMTT | No | Yes | Webserver Source code | [4] |
CCTop | University of Heidelberg | Yes | Yes | Partial | 5 (0-5) | No | NGG, NRG, NNGRRT, NNNNGATT, NNAGAAW, NAAAAC | Yes | Yes | Webserver | [5] |
CHOPCHOP | Harvard University | Yes | Yes | Partial | 0, 2 | No | NGG, NNAGAA, NNNNGANN | No | Yes | Webserver | [6] |
CHOPCHOP v2 | University of Bergen | Yes | Yes | Yes | 3 (0-3) | Yes | User customizable | Yes | Yes | Webserver | [7] |
COD | Dayong Guo | No | No | No | 0, 3, 5, 8 | No | NGG and NAG | No | Yes | Webserver | - |
CRISPR Configurator & Specificity Tool | Dharmacon, Inc. | Yes (over 30 species) | Yes | Yes | 8 (gaps or mismatches) | Internally | NGG and NAG | mRNA exons, Links to UCSC genome browser annotations | No | Webserver Specificity Tool | - |
CRISPR Design | Zhang Lab, MIT | Yes | No | No | 4 | No | NGG and NAG | mRNA exons | Yes | Webserver | [8] |
CRISPRdirect | Database Center for Life Science (DBCLS) | Yes (over 200 species) | Yes | No | Any number | No | NNN | Yes | Yes | Webserver | [9] |
CRISPR gRNA Design Tool | DNA2.0 | Yes | Yes | No | 0-10 | No | NGG, NAG | Genbank annotations: Gene, misc_RNA, ncRNA, CDS, exon | Yes | Webserver | - |
CRISPRseek | Bioconductor | Yes | Yes | No | Any number | No | User customizable | mRNA exons | Yes | Source code | [10] |
DESKGEN | Desktop Genetics | Yes | Yes | Yes | Any number | Yes | Fully user customizable | Yes | Yes | Webserver | [11] |
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 | Webserver Source code Remote batch submission | [12] | |
sgRNA Designer | Broad Institute | No | No | No | 0 | Yes | NGG | CDS (if searching by transcript ID) | Yes | Webserver Source code | [1] |
References
- 1 2 3 Doench JG, Hartenian E, Graham DB, Tothova Z, Hegde M, Smith I, Sullender M, Ebert BL, Xavier RJ, Root DE (2014). "Rational design of highly active sgRNAs for CRISPR-Cas9-mediated gene inactivation". Nat. Biotechnol. 32 (12): 1262–7. doi:10.1038/nbt.3026. PMID 25184501.
- 1 2 Chari, Raj; Mali, Prashant; Moosburner, Mark; Church, George M (2015-01-01). "Unraveling CRISPR-Cas9 genome engineering parameters via a library-on-library approach". Nature Methods. 12: 823–826. doi:10.1038/nmeth.3473.
- ↑ Oliveros, Juan C.; Franch, Mònica; Tabas-Madrid, Daniel; San-León, David; Montoliu, Lluis; Cubas, Pilar; Pazos, Florencio (2016-07-08). "Breaking-Cas-interactive design of guide RNAs for CRISPR-Cas experiments for ENSEMBL genomes". Nucleic Acids Research. 44 (W1): W267–271. doi:10.1093/nar/gkw407. ISSN 1362-4962. PMC 4987939
. PMID 27166368.
- ↑ Bae S, Park J, Kim JS (2014). "Cas-OFFinder: a fast and versatile algorithm that searches for potential off-target sites of Cas9 RNA-guided endonucleases". Bioinformatics. 30 (10): 1473–5. doi:10.1093/bioinformatics/btu048. PMID 24463181.
- ↑ Stemmer M, Thumberger T, Del Sol Keyer M, Wittbrodt J, Mateo JL (2015). "CCTop: An Intuitive, Flexible and Reliable CRISPR/Cas9 Target Prediction Tool". PLOS ONE. 10: e0124633. doi:10.1371/journal.pone.0124633. PMC 4409221
. PMID 25909470.
- ↑ Montague TG, Cruz JM, Gagnon JA, Church GM, Valen E (2014). "CHOPCHOP: a CRISPR/Cas9 and TALEN web tool for genome editing". Nucleic Acids Res. 42 (W1): W401–7. doi:10.1093/nar/gku410. PMID 24861617.
- ↑ Labun K, Montague TG, Gagnon JA, Thyme SB, Valen E (2016). "CHOPCHOP v2: a web tool for the next generation of CRISPR genome engineering.". Nucleic Acids Res. 44 (W1): W272–6. doi:10.1093/nar/gkw398. PMC 4987937
. PMID 27185894.
- ↑ 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 (2013). "DNA targeting specificity of RNA-guided Cas9 nucleases". Nat. Biotechnol. 31 (9): 827–32. doi:10.1038/nbt.2647. PMID 23873081.
- ↑ Naito Y, Hino K, Bono H, Ui-Tei K (2015). "CRISPRdirect: software for designing CRISPR/Cas guide RNA with reduced off-target sites". Bioinformatics. 31 (7): 1120–3. doi:10.1093/bioinformatics/btu743. PMC 4382898
. PMID 25414360.
- ↑ Zhu LJ, Holmes BR, Aronin N, Brodsky MH (2014). "CRISPRseek: A Bioconductor Package to Identify Target-Specific Guide RNAs for CRISPR-Cas9 Genome-Editing Systems". PLOS ONE. 9 (9): e108424. doi:10.1371/journal.pone.0108424. PMC 4172692
. PMID 25247697.
- ↑ "Desktop Genetics Announces the Launch of DeskGen Gene Editing Platform". American Laboratory. 2015.
- ↑ Pliatsika V, Rigoutsos I (2015). ""Off-Spotter": very fast and exhaustive enumeration of genomic lookalikes for designing CRISPR/ Cas guide RNAs". Biol. Direct. 10 (1): 4. doi:10.1186/s13062-015-0035-z. PMID 25630343.
External links
- Off-Spotter
- Benchling CRISPR
- Breaking-Cas
- CRISPRseek
- Cas-OFFinder
- DESKGEN
- MIT CRISPR Design
- DNA2.0 CRISPR gRNA design tool
- CCtop
- ChopChop
- ChopChop v2
- ChopChop script version
- Doench sgRNA designer
- CRISPy for CHO-K1
- Dharmacon CRISPR RNA Configurator
- Dharmacon CRISPR Specificity Tool
- CRISPRdirect