Multi-locus CRISPRi targeting with a single truncated guide RNA

Molly M. Moore, Siddarth Wekhande, Robbyn Issner, Alejandro Collins, Anna J. Cruz, Yanjing V. Liu, Nauman Javed, Salvador Casaní-Galdón, Jason D. Buenrostro, Charles B. Epstein, Eugenio Mattei, John G. Doench, Bradley E. Bernstein, Noam Shoresh and Fadi J. Najm ()
Additional contact information
Molly M. Moore: Broad Institute of MIT and Harvard
Siddarth Wekhande: Broad Institute of MIT and Harvard
Robbyn Issner: Broad Institute of MIT and Harvard
Alejandro Collins: Broad Institute of MIT and Harvard
Anna J. Cruz: Broad Institute of MIT and Harvard
Yanjing V. Liu: Broad Institute of MIT and Harvard
Nauman Javed: Broad Institute of MIT and Harvard
Salvador Casaní-Galdón: Broad Institute of MIT and Harvard
Jason D. Buenrostro: Broad Institute of MIT and Harvard
Charles B. Epstein: Broad Institute of MIT and Harvard
Eugenio Mattei: Broad Institute of MIT and Harvard
John G. Doench: Broad Institute of MIT and Harvard
Bradley E. Bernstein: Broad Institute of MIT and Harvard
Noam Shoresh: Broad Institute of MIT and Harvard
Fadi J. Najm: Broad Institute of MIT and Harvard

Nature Communications, 2025, vol. 16, issue 1, 1-12

Abstract: Abstract A critical goal in functional genomics is evaluating which non-coding elements contribute to gene expression, cellular function, and disease. Functional characterization remains a challenge due to the abundance and complexity of candidate elements. Here, we develop a CRISPRi-based approach for multi-locus screening of putative transcription factor binding sites with a single truncated guide. A truncated guide with hundreds of sequence match sites can reliably disrupt enhancer activity, which expands the targeting scope of CRISPRi while maintaining repressive efficacy. We screen over 13,000 possible CTCF binding sites with 24 guides at 10 nucleotides in spacer length. These truncated guides direct CRISPRi-mediated deposition of repressive H3K9me3 marks and disrupt transcription factor binding at most sequence match target sites. This approach can be a valuable screening step for testing transcription factor binding motifs or other repeated genomic sequences and is easily implemented with existing tools.

Date: 2025
References: View references in EconPapers View complete reference list from CitEc
Citations:

Downloads: (external link)
https://www.nature.com/articles/s41467-025-56144-x Abstract (text/html)

Related works:
This item may be available elsewhere in EconPapers: Search for items with the same title.

Export reference: BibTeX RIS (EndNote, ProCite, RefMan) HTML/Text

Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:16:y:2025:i:1:d:10.1038_s41467-025-56144-x

Ordering information: This journal article can be ordered from
https://www.nature.com/ncomms/

DOI: 10.1038/s41467-025-56144-x

Access Statistics for this article

Nature Communications is currently edited by Nathalie Le Bot, Enda Bergin and Fiona Gillespie

More articles in Nature Communications from Nature
Bibliographic data for series maintained by Sonal Shukla () and Springer Nature Abstracting and Indexing ().