sgRNAcas9: a software package for designing CRISPR sgRNA and evaluating potential off-target cleavage sites

Although the CRISPR/Cas9/sgRNA system efficiently cleaves intracellular DNA at desired target sites, major concerns remain on potential "off-target" cleavage that may occur throughout the whole genome. In order to improve CRISPR-Cas9 specificity for targeted genome editing and transcriptio...

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Bibliographic Details
Published in:PloS one 2014-06, Vol.9 (6), p.e100448-e100448
Main Authors: Xie, Shengsong, Shen, Bin, Zhang, Chaobao, Huang, Xingxu, Zhang, Yonglian
Format: Article
Language:English
Subjects:
Algorithms
Applications programs
Base Sequence
Biochemistry
Bioinformatics
Biology
Biology and Life Sciences
Cleavage
Clustered Regularly Interspaced Short Palindromic Repeats - genetics
Computational Biology - methods
Computer and Information Sciences
Computer programs
CRISPR
Deoxyribonucleic acid
Design
DNA
DNA Cleavage
Editing
Expression vectors
Gene expression
Genetic disorders
Genetic engineering
Genome editing
Genomes
Genomics
Humans
Laboratories
Molecular Sequence Data
Mutation
Nucleotides
Oligonucleotides
Operating systems
RNA Editing - genetics
RNA, Guide, CRISPR-Cas Systems - genetics
Sequence Homology, Nucleic Acid
Software
Software industry
Software packages
Transcription
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Summary:Although the CRISPR/Cas9/sgRNA system efficiently cleaves intracellular DNA at desired target sites, major concerns remain on potential "off-target" cleavage that may occur throughout the whole genome. In order to improve CRISPR-Cas9 specificity for targeted genome editing and transcriptional control, we describe a bioinformatics tool "sgRNAcas9", which is a software package developed for fast design of CRISPR sgRNA with minimized off-target effects. This package consists of programs to perform a search for CRISPR target sites (protospacers) with user-defined parameters, predict genome-wide Cas9 potential off-target cleavage sites (POT), classify the POT into three categories, batch-design oligonucleotides for constructing 20-nt (nucleotides) or truncated sgRNA expression vectors, extract desired length nucleotide sequences flanking the on- or off-target cleavage sites for designing PCR primer pairs to validate the mutations by T7E1 cleavage assay. Importantly, by identifying potential off-target sites in silico, the sgRNAcas9 allows the selection of more specific target sites and aids the identification of bona fide off-target sites, significantly facilitating the design of sgRNA for genome editing applications. sgRNAcas9 software package is publicly available at BiooTools website (www.biootools.com) under the terms of the GNU General Public License.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0100448