Rxivist logo

Optimized RNA-targeting CRISPR/Cas13d technology outperforms shRNA in identifying essential circRNAs

By Yang Zhang, Tuan M. Nguyen, Xiao-Ou Zhang, Tin Phan, John G. Clohessy, Pier Paolo Pandolfi

Posted 23 Mar 2020
bioRxiv DOI: 10.1101/2020.03.23.002238

Circular RNAs (circRNAs) are widely expressed, but their functions remain largely unknown. To study circRNAs in a high-throughput manner, short hairpin RNA (shRNA) screens have recently been used to deplete circRNAs by targeting their unique back-splicing junction (BSJ) sites. Here, we report frequent discrepancies between shRNA-mediated circRNA knockdown efficiency and the corresponding biological effect, raising pressing concerns about the robustness of shRNA screening for functional circRNAs. To address this issue, we leveraged the CRISPR/Cas13d system for circRNAs functional screenings. We optimized a strategy for designing single guide RNAs to deplete circRNAs. We then performed shRNA and CRISPR/Cas13d parallel screenings and demonstrated that shRNA-mediated circRNAs screening yielded a high rate of false positives phenotypes, while optimized CRISPR/Cas13d led to the identification of bona-fide functional circRNAs. Collectively, we developed a specific and reliable approach to functionalize circRNAs in a high-throughput manner.

Download data

  • Downloaded 1,147 times
  • Download rankings, all-time:
    • Site-wide: 31,111
    • In molecular biology: 792
  • Year to date:
    • Site-wide: 91,491
  • Since beginning of last month:
    • Site-wide: 105,379

Altmetric data


Downloads over time

Distribution of downloads per paper, site-wide


PanLingua

News