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

Publication ,  Journal Article
Ousterout, DG; Kabadi, AM; Thakore, PI; Majoros, WH; Reddy, TE; Gersbach, CA
Published in: Nat Commun
February 18, 2015

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.

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Published In

Nat Commun

DOI

EISSN

2041-1723

Publication Date

February 18, 2015

Volume

6

Start / End Page

6244

Location

England

Related Subject Headings

  • Polymerase Chain Reaction
  • Plasmids
  • Mutation
  • Muscular Dystrophy, Duchenne
  • Mice, SCID
  • Mice
  • Male
  • Humans
  • High-Throughput Nucleotide Sequencing
  • HEK293 Cells
 

Citation

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Ousterout, D. G., Kabadi, A. M., Thakore, P. I., Majoros, W. H., Reddy, T. E., & Gersbach, C. A. (2015). Multiplex CRISPR/Cas9-based genome editing for correction of dystrophin mutations that cause Duchenne muscular dystrophy. Nat Commun, 6, 6244. https://doi.org/10.1038/ncomms7244
Ousterout, David G., Ami M. Kabadi, Pratiksha I. Thakore, William H. Majoros, Timothy E. Reddy, and Charles A. Gersbach. “Multiplex CRISPR/Cas9-based genome editing for correction of dystrophin mutations that cause Duchenne muscular dystrophy.Nat Commun 6 (February 18, 2015): 6244. https://doi.org/10.1038/ncomms7244.
Ousterout DG, Kabadi AM, Thakore PI, Majoros WH, Reddy TE, Gersbach CA. Multiplex CRISPR/Cas9-based genome editing for correction of dystrophin mutations that cause Duchenne muscular dystrophy. Nat Commun. 2015 Feb 18;6:6244.
Ousterout, David G., et al. “Multiplex CRISPR/Cas9-based genome editing for correction of dystrophin mutations that cause Duchenne muscular dystrophy.Nat Commun, vol. 6, Feb. 2015, p. 6244. Pubmed, doi:10.1038/ncomms7244.
Ousterout DG, Kabadi AM, Thakore PI, Majoros WH, Reddy TE, Gersbach CA. Multiplex CRISPR/Cas9-based genome editing for correction of dystrophin mutations that cause Duchenne muscular dystrophy. Nat Commun. 2015 Feb 18;6:6244.

Published In

Nat Commun

DOI

EISSN

2041-1723

Publication Date

February 18, 2015

Volume

6

Start / End Page

6244

Location

England

Related Subject Headings

  • Polymerase Chain Reaction
  • Plasmids
  • Mutation
  • Muscular Dystrophy, Duchenne
  • Mice, SCID
  • Mice
  • Male
  • Humans
  • High-Throughput Nucleotide Sequencing
  • HEK293 Cells