Table 1.
Commonly used sgRNA design tools and databases
| Name | Organism | Cas nuclease | Major feature | Database or web server | Website | Refs. |
|---|---|---|---|---|---|---|
| CRISPOR | > 100 species | > 30 Cas9 orthologues and Cas variants | Designing, evaluating, and cloning guide sequences for the CRISPR/Cas9 system; providing primers for vector construction; indicating mismatch number; and linking off-target to genome browser | Web server | https://crispor.tefor.net/ | [38] |
| CHOPCHOP | > 100 species | Cas9, Cas12, Cas13, and TALEN | Providing multiple predictive models; visualizing genomic location of targets and genes; and providing primers | Web server | https://chopchop.cbu.uib.no/ | [46], [47] |
| CRISPR RGEN Tools | > 100 species | > 20 Cas9 orthologues and Cas variants | Providing multiple predictive models; downloadable and standalone; and predicting potential off-target number via Cas-OFFinder, and out-of-frame scores via Microhomology-Predictor | Web server | https://www.rgenome.net/cas-designer/ | [84], [93] |
| E-CRISP | > 50 species | SpCas9 | Feasibly creating genome-scale libraries; downloadable; and frequently updated | Web server | https://www.e-crisp.org/E-CRISP/index.html | [49] |
| GUIDES | Human and mouse | SpCas9 | Feasibly designing CRISPR knockout libraries; downloadable; and step-by-step | Web server | https://guides.sanjanalab.org/ and https://github.com/sanjanalab/GUIDES | [11] |
| CRISPRscan | > 10 species | Cas9 and Cas12 | Designing sgRNAs for protein-coding genes; ready-to-inject sgRNA sequence; tracks for genome browser; and searching whole-genome off-target impacts | Web server | https://www.crisprscan.org/ | [45] |
| CCTop | > 100 species | > 10 Cas9 orthologues and Cas variants | Searching for single and multiple queries; indicating mismatch number; predicting off-target impacts; and predicting sgRNA efficiency using CRISPRater with custom in vitro transcription selection | Web server | https://cctop.cos.uni-heidelberg.de/ | [39] |
| CRISTA | > 100 species | SpCas9 | Providing machine learning framework, including DNA/RNA bulge genomic context and RNA thermodynamics; detecting off-targets; and ranking targets | Web server | https://crista.tau.ac.il/ | [56] |
| DeepCRISPR | Human | SpCas9 | Incorporating epigenetic information; and predicting off-target impacts | Web server | https://www.deepcrispr.net/ | [57] |
| DRSC Find CRISPRs | Drosophila | SpCas9 | Providing off-target stringency from 3 to 5 mismatches; and separating target region and potential off-targets by different tracts | Web server |
https://www.flyrnai.org/crispr/ https://www.flyrnai.org/crispr3/web/ |
[72] |
| EuPaGDT | Eukaryotic pathogens | > 10 Cas9 orthologues and Cas variants | Providing wide compatibility for eukaryotic pathogen genomes | Web server | https://grna.ctegd.uga.edu/ | [73] |
| WU-CRISPR | Human and mouse | SpCas9 | Providing machine learning algorithm trained by experimental data; providing custom sequence between 26 bp and 30,000 bp with one sequence per time; and downloadable results | Web server | https://crispr.wustl.edu/ | [155], [156] |
| GPP sgRNA Designer | Human, mouse, and rat | SpCas9, SaCas9, and AsCpf1 | Inputting up to 200 transcript IDs or gene IDs; maximizing on-target activity and minimizing off-target activity; and scoring on-targeting efforts | Web server | https://portals.broadinstitute.org/gpp/public/analysis-tools/sgrna-design | [48] |
| CRISPR-GE | > 40 plant species | SpCas9, FnCpf1, and AsCpf1 | Providing software toolkits, primer design for vector construction, on-target amplification, and PCR sequencing result analysis | Web server | https://skl.scau.edu.cn/ | [94] |
| CRISPR-P | 49 plant species | > 14 Cas9 and variants | Supporting wide range of plant species; providing on-target and off-target scoring; and providing gRNA sequence analysis | Web server | https://crispr.hzau.edu.cn/CRISPR2/ | [95], [96] |
| CRISPR-PLANT V2 | 7 plant species | SpCas9 | Supporting main model and crop plant species; providing selection of chromosome and locations with clear instruction | Web server | https://www.genome.arizona.edu/crispr2/ | [157] |
| CRISPRz | Zebrafish, human, and mouse | SpCas9 | Providing specific for a wide variety of cell lines and organisms including zebrafish; and providing validated sgRNA database | Web server | https://research.nhgri.nih.gov/CRISPRz/ | [82] |
| CRISPRlnc | 10 species | SpCas9 | Providing downloadable validated sgRNA database for lncRNAs | Database | https://www.crisprlnc.org/ | [81] |
| FORECasT | Human | SpCas9 | Predicting the mutational outcomes | Web server | https://partslab.sanger.ac.uk/FORECasT | [87] |
| AsCRISPR | Human and mouse | SpCas9, AsCpf1, AaCas12b, CasX, and variants | Designing sgRNAs for allele-specific genetic elements | Web server | https://www.genemed.tech/ascrispr/ascrispr | [98] |
| SNP-CRISPR | 9 plant and animal species | NGG and NAG PAM | Designing sgRNAs for targeting SNPs or Indel variants | Web server | https://www.flyrnai.org/tools/snp_crispr/web/ | [99] |
| SSC | N/A | Cas9 | For both CRISPR knockout and CRISPRa/CRISPRi | Web server | https://cistrome.org/SSC/ | [35] |
| DeepHF | N/A | SPCas9 and Cas9HF | gRNA designer and efficiency prediction | Web server and database | https://www.DeepHF.com/ | [158] |
| PnB Designer | 6 species | Cas9 | Designing pegRNAs for prime editors and sgRNAs for base editors | Web server | https://fgcz-shiny.uzh.ch/PnBDesigner/ | [99], [100] |
| inDelphi | Human | SpCas9 | Predicting the mutational outcomes | Web server | https://www.crisprindelphi.design/ | [86] |
Note: Cas, CRISPR-associated protein; CRISPR, clustered regularly interspaced short palindromic repeats; CRISPRa, CRISPR activation; CRISPRi, CRISPR interference; gRNA, guide RNA; lncRNA, long non-coding RNA; pegRNA; prime editing guide RNA; sgRNA, single guide RNA; TALEN, transcription activator-like effector nuclease; N/A, not available.