CRISPR-COPIES: an in silico platform for discovery of neutral integration sites for CRISPR/Cas-facilitated gene integration

Boob, Aashutosh Girish; Zhu, Zhixin; Intasian, Pattarawan; Jain, Manan; Petrov, Vassily Andrew; Lane, Stephan Thomas; Tan, Shih-I; Xun, Guanhua; Zhao, Huimin

doi:10.1093/nar/gkae062

Title: CRISPR-COPIES: an in silico platform for discovery of neutral integration sites for CRISPR/Cas-facilitated gene integration

Journal Article · 12 February 2024 · Nucleic Acids Research

DOI: https://doi.org/10.1093/nar/gkae062 · OSTI ID:2301778

Boob, Aashutosh Girish; Zhu, Zhixin; Intasian, Pattarawan; Jain, Manan; Petrov, Vassily Andrew; Lane, Stephan Thomas; Tan, Shih-I; Xun, Guanhua;

Abstract The CRISPR/Cas system has emerged as a powerful tool for genome editing in metabolic engineering and human gene therapy. However, locating the optimal site on the chromosome to integrate heterologous genes using the CRISPR/Cas system remains an open question. Selecting a suitable site for gene integration involves considering multiple complex criteria, including factors related to CRISPR/Cas-mediated integration, genetic stability, and gene expression. Consequently, identifying such sites on specific or different chromosomal locations typically requires extensive characterization efforts. To address these challenges, we have developed CRISPR-COPIES, a COmputational Pipeline for the Identification of CRISPR/Cas-facilitated intEgration Sites. This tool leverages ScaNN, a state-of-the-art model on the embedding-based nearest neighbor search for fast and accurate off-target search, and can identify genome-wide intergenic sites for most bacterial and fungal genomes within minutes. As a proof of concept, we utilized CRISPR-COPIES to characterize neutral integration sites in three diverse species: Saccharomyces cerevisiae, Cupriavidus necator, and HEK293T cells. In addition, we developed a user-friendly web interface for CRISPR-COPIES (https://biofoundry.web.illinois.edu/copies/). We anticipate that CRISPR-COPIES will serve as a valuable tool for targeted DNA integration and aid in the characterization of synthetic biology toolkits, enable rapid strain construction to produce valuable biochemicals, and support human gene and cell therapy applications.

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Research Organization:: Center for Advanced Bioenergy and Bioproducts Innovation (CABBI), Urbana, IL (United States); Univ. of Illinois at Urbana-Champaign, IL (United States)

Sponsoring Organization:: USDOE; USDOE Office of Science (SC), Biological and Environmental Research (BER)

Grant/Contract Number:: SC0018420

OSTI ID:: 2301778

Journal Information:: Nucleic Acids Research, Journal Name: Nucleic Acids Research Journal Issue: 6 Vol. 52; ISSN 0305-1048

Publisher:: Oxford University PressCopyright Statement

Country of Publication:: United Kingdom

Language:: English

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Title: CRISPR-COPIES: an in silico platform for discovery of neutral integration sites for CRISPR/Cas-facilitated gene integration

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