Efficient Recreation of T(11;22) EWSR1-FLI1 in Human Stem Cells Using CRISPR/Cas9

Overview

Journal Stem Cell Reports

Publisher Cell Press

Specialty Cell Biology

Date 2017 May 13

PMID 28494941

Citations 33

Authors

Raul Torres-Ruiz

Marta Martinez-Lage

Maria C Martin

Aida Garcia

Clara Bueno

Julio Castano

Juan C Ramirez

Pablo Menendez

Juan C Cigudosa

Sandra Rodriguez-Perales

Affiliations

Soon will be listed here.

Abstract

Efficient methodologies for recreating cancer-associated chromosome translocations are in high demand as tools for investigating how such events initiate cancer. The CRISPR/Cas9 system has been used to reconstruct the genetics of these complex rearrangements at native loci while maintaining the architecture and regulatory elements. However, the CRISPR system remains inefficient in human stem cells. Here, we compared three strategies aimed at enhancing the efficiency of the CRISPR-mediated t(11;22) translocation in human stem cells, including mesenchymal and induced pluripotent stem cells: (1) using end-joining DNA processing factors involved in repair mechanisms, or (2) ssODNs to guide the ligation of the double-strand break ends generated by CRISPR/Cas9; and (3) all-in-one plasmid or ribonucleoprotein complex-based approaches. We report that the generation of targeted t(11;22) is significantly increased by using a combination of ribonucleoprotein complexes and ssODNs. The CRISPR/Cas9-mediated generation of targeted t(11;22) in human stem cells opens up new avenues in modeling Ewing sarcoma.

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