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CRISPR-Cas9 has quickly become the method of choice for genome editing, with multiple publications describing technical advances and novel applications. It has been widely adopted as a tool for basic research and has significant translational and clinical potential. However, its usage has outpaced the establishment of essential and rigorous controls for unwanted off-target effects, manifested as small mutations, large deletions of target loci, or large-scale chromosomal rearrangements. A common application of CRISPR-Cas9 is as a tool for creating isogenic cell-line models to study the effects of precise mutations, or variants, on disease traits. Here, we describe the effect of standard CRISPR-Cas9 mutagenesis protocols on well characterized cancer cell lines. We demonstrate that commonly used methods for detecting correctly mutated clones fail to uncover large-scale rearrangements. We show that simple cytogenetic methods can be used to identify clones carrying chromosomal abnormalities and large mutations at target loci. These methods are quick and cost-efficient, and we suggest that such controls should be performed prior to publication of studies based on novel CRISPR-Cas9 mutated cancer cell lines.

Original publication

DOI

10.1089/crispr.2019.0006

Type

Journal article

Journal

CRISPR J

Publication Date

12/2019

Volume

2

Pages

406 - 416

Keywords

CRISPR-Cas Systems, Cell Line, Tumor, Chromosomal Instability, Clustered Regularly Interspaced Short Palindromic Repeats, Cytogenetic Analysis, Gene Editing, Gene Rearrangement, Humans, Mutagenesis, Mutation, Neoplasms, RNA, Guide, Kinetoplastida, Sequence Deletion