Rxivist logo

Rxivist combines preprints from bioRxiv with data from Twitter to help you find the papers being discussed in your field. Currently indexing 72,942 bioRxiv papers from 317,562 authors.

Pooled CRISPR-Cas9 screens have recently emerged as a powerful method for functionally characterizing regulatory elements in the non-coding genome, but off-target effects in these experiments have not been systematically evaluated. Here, we conducted a genome-scale screen for essential CTCF loop anchors in the K562 leukemia cell line. Surprisingly, the primary drivers of signal in this screen were single guide RNAs (sgRNAs) with low specificity scores. After removing these guides, we found that there were no CTCF loop anchors critical for cell growth. We also observed this effect in an independent screen fine-mapping the core motifs in enhancers of the GATA1 gene. We then conducted screens in parallel with CRISPRi and CRISPRa, which do not induce DNA damage, and found that an unexpected and distinct set of off-targets also caused strong confounding growth effects with these epigenome-editing platforms. Promisingly, strict filtering of CRISPRi libraries using GuideScan specificity scores removed these confounded sgRNAs and allowed for the identification of essential enhancers, which we validated extensively. Together, our results show off-target activity can severely limit identification of essential functional motifs by active Cas9, while strictly filtered CRISPRi screens can be reliably used for assaying larger regulatory elements.

Download data

  • Downloaded 1,753 times
  • Download rankings, all-time:
    • Site-wide: 3,346 out of 72,919
    • In genomics: 640 out of 4,843
  • Year to date:
    • Site-wide: 7,577 out of 72,919
  • Since beginning of last month:
    • Site-wide: 7,577 out of 72,919

Altmetric data


Downloads over time

Distribution of downloads per paper, site-wide


PanLingua

Sign up for the Rxivist weekly newsletter! (Click here for more details.)


News