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Nat Commun. 2015 Feb 18;6:6244. doi: 10.1038/ncomms7244.

Multiplex CRISPR/Cas9-based genome editing for correction of dystrophin mutations that cause Duchenne muscular dystrophy.

Author information

1
Department of Biomedical Engineering, Duke University, Room 136 Hudson Hall, Box 90281, Durham, North Carolina 27708, USA.
2
Program in Computational Biology and Bioinformatics, Duke University, Durham, North Carolina 27708, USA.
3
1] Center for Genomic and Computational Biology, Duke University, Durham, North Carolina 27708, USA [2] Department of Biostatistics and Bioinformatics, Duke University Medical Center, Durham, North Carolina 27710, USA.
4
1] Department of Biomedical Engineering, Duke University, Room 136 Hudson Hall, Box 90281, Durham, North Carolina 27708, USA [2] Center for Genomic and Computational Biology, Duke University, Durham, North Carolina 27708, USA [3] Department of Orthopaedic Surgery, Duke University Medical Center, Durham, North Carolina 27710, USA.

Abstract

The CRISPR/Cas9 genome-editing platform is a promising technology to correct the genetic basis of hereditary diseases. The versatility, efficiency and multiplexing capabilities of the CRISPR/Cas9 system enable a variety of otherwise challenging gene correction strategies. Here, we use the CRISPR/Cas9 system to restore the expression of the dystrophin gene in cells carrying dystrophin mutations that cause Duchenne muscular dystrophy (DMD). We design single or multiplexed sgRNAs to restore the dystrophin reading frame by targeting the mutational hotspot at exons 45-55 and introducing shifts within exons or deleting one or more exons. Following gene editing in DMD patient myoblasts, dystrophin expression is restored in vitro. Human dystrophin is also detected in vivo after transplantation of genetically corrected patient cells into immunodeficient mice. Importantly, the unique multiplex gene-editing capabilities of the CRISPR/Cas9 system facilitate the generation of a single large deletion that can correct up to 62% of DMD mutations.

PMID:
25692716
PMCID:
PMC4335351
DOI:
10.1038/ncomms7244
[Indexed for MEDLINE]
Free PMC Article

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