CRISPR Guide RNA Design Guidelines for Efficient Genome Editing

Patrick Schindele; Felix Wolter; Holger Puchta

doi:10.1007/978-1-0716-0712-1_19

CRISPR Guide RNA Design Guidelines for Efficient Genome Editing

Methods Mol Biol. 2020:2166:331-342. doi: 10.1007/978-1-0716-0712-1_19.

Authors

Patrick Schindele¹, Felix Wolter¹, Holger Puchta²

Affiliations

¹ Botanical Institute, Karlsruhe Institute of Technology, Karlsruhe, Germany.
² Botanical Institute, Karlsruhe Institute of Technology, Karlsruhe, Germany. holger.puchta@kit.edu.

PMID: 32710418
DOI: 10.1007/978-1-0716-0712-1_19

Abstract

The simple applicability and facile target programming of the CRISPR/Cas9-system abolish the major boundaries of previous genome editing tools, making it the tool of choice for generating site-specific genome alterations. Its versatility and efficacy have been demonstrated in various organisms; however, accurately predicting guide RNA efficiencies remains an organism-independent challenge. Thus, designing optimal guide RNAs is essential to maximize the experimental outcome. Here, we summarize the current knowledge for guide RNA design and highlight discrepancies between different experimental systems.

Keywords: CRISPR; CRISPR prediction tool; Cas9; Genome editing; Mismatch tolerance; gRNA design; gRNA secondary structure.

MeSH terms

Animals
CRISPR-Associated Protein 9 / genetics
CRISPR-Cas Systems*
Chromatin / genetics
Clustered Regularly Interspaced Short Palindromic Repeats / genetics*
Drosophila / genetics
Gene Editing / methods*
Genome
HEK293 Cells
Humans
Nucleic Acid Conformation
Nucleotides / chemistry
RNA, Guide, CRISPR-Cas Systems / chemistry
RNA, Guide, CRISPR-Cas Systems / genetics*
Streptococcus pyogenes / enzymology

Substances

Chromatin
Nucleotides
RNA, Guide, CRISPR-Cas Systems
CRISPR-Associated Protein 9