
CRISPR-based CTRL-Mutagenesis induces diverse mutations in target regions for functional analysis of non-coding elements.


coremsg

Functional regions that regulate biological phenomena are interspersed throughout eukaryotic genomes. The most definitive approach for identifying such regions is to confirm the phenotype of cells or organisms in which specific regions have been mutated or removed from the genome. This approach is invaluable for the functional analysis of genes with a defined functional element, the protein-coding sequence. In contrast, no functional analysis platforms have been established for the study of cis-elements or microRNA cluster regions consisting of multiple microRNAs with functional overlap. Whole-genome mutagenesis approaches, such as via N -ethyl- N -nitrosourea (ENU) and gene trapping, have greatly contributed to elucidating the function of coding genes. These methods almost never induce deletions of genomic regions or multiple mutations within a narrow region. In other words, cis-elements and microRNA clusters cannot be effectively targeted in such a manner. Herein, we established a novel region-specific random mutagenesis method named CRISPR-& Transposase-based RegionaL Mutagenesis (CTRL-Mutagenesis). We demonstrated that CTRL-mutagenesis randomly induces diverse mutations within target regions in murine embryonic stem cells. Comparative analysis of mutants harbouring subtly different mutations within the same region would facilitate the further study of cis-element and microRNA clusters.

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### Competing Interest Statement

The authors have declared no competing interest.

 [1]: pending:yes

Regional random mutagenesis driven by multiple sgRNAs and diverse on-target genome editing events to identify functionally important elements in non-coding regions

regional-random-mutagenesis-for-functional-elements-identification


Regional Random Mutagenesis for Functional Elements Identification
