Journal
GENOME BIOLOGY
Volume 22, Issue 1, Pages -Publisher
BMC
DOI: 10.1186/s13059-021-02263-9
Keywords
circRNAs; CRISPR; Cas13d system; High-throughput screening
Funding
- R35 NCI [CA197529-01]
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By using the CRISPR/Cas13d tool and optimizing the design strategy, the specificity of circRNA silencing is significantly enhanced and validated in parallel screenings. Utilizing CRISPR/Cas13d for high-throughput study of functional circRNAs proves to be an effective approach.
Short hairpin RNAs (shRNAs) are used to deplete circRNAs by targeting back-splicing junction (BSJ) sites. However, frequent discrepancies exist between shRNA-mediated circRNA knockdown and the corresponding biological effect, querying their robustness. By leveraging CRISPR/Cas13d tool and optimizing the strategy for designing single-guide RNAs against circRNA BSJ sites, we markedly enhance specificity of circRNA silencing. This specificity is validated in parallel screenings by shRNA and CRISPR/Cas13d libraries. Using a CRISPR/Cas13d screening library targeting > 2500 human hepatocellular carcinoma-related circRNAs, we subsequently identify a subset of sorafenib-resistant circRNAs. Thus, CRISPR/Cas13d represents an effective approach for high-throughput study of functional circRNAs.
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