Application of CRISPR/cas9-Directed Homologous Recombination to the Generation of Human Tumor Cells with Conditional Knockout of an X-Linked MicroRNA Locus

Aurélie Van Tongelen, Axelle Loriot, Olivier De Backer, Charles De Smet

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Studying the cellular function of microRNAs requires genetic strategies to generate their loss-of-function. Recently, a novel approach of targeted genomic deletion was proposed, which is based on induction of site-specific DNA cuts with Cas9/gRNA ribonucleoprotein complexes, combined with homologous recombination-dependent insertion of cassettes that contain sequences for Cre-lox or FLP-FRT systems. Here, we provide a technical report describing application of this CRISPR/Cas9-directed homologous recombination procedure to the generation of human tumor cells in which conditional knockout of an X-linked cluster of microRNAs (miR-105/miR-767) can be induced. We describe the successive steps of genetic engineering and cell clone selection that allowed us to generate cells with the expected genome editing.
Original languageEnglish
JournalGene Technology
Issue number4:124
Publication statusPublished - 20 Jul 2015

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