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Functional repair of a mutant chloride channel using a trans-splicing ribozyme.

Publication ,  Journal Article
Rogers, CS; Vanoye, CG; Sullenger, BA; George, AL
Published in: J Clin Invest
December 2002

RNA repair has been proposed as a novel gene-based therapeutic strategy. Modified Tetrahymena group I intron ribozymes have been used to mediate trans-splicing of therapeutically relevant RNA transcripts, but the efficiency of the ribozyme-mediated RNA repair process has not been determined precisely and subsequent restoration of protein function has been demonstrated only by indirect means. We engineered a ribozyme that targets the mRNA of a mutant canine skeletal muscle chloride channel (cClC-1) (mutation T268M in ClC-1 causing myotonia congenita) and replaces the mutant-containing 3' portion by trans-splicing the corresponding 4-kb wild-type sequence. Repair efficiency assessed by quantitative RT-PCR was 1.2% +/- 0.1% in a population of treated cells. However, when chloride channel function was examined in single cells, a wide range of electrophysiological activity was observed, with 18% of cells exhibiting significant functional restoration and some cells exhibiting complete rescue of the biophysical phenotype. These results indicate that RNA repair can restore wild-type protein activity and reveal considerable cell-to-cell variability in ribozyme-mediated trans-splicing reaction efficiency.

Duke Scholars

Published In

J Clin Invest

DOI

ISSN

0021-9738

Publication Date

December 2002

Volume

110

Issue

12

Start / End Page

1783 / 1789

Location

United States

Related Subject Headings

  • Trans-Splicing
  • Tetrahymena
  • Reverse Transcriptase Polymerase Chain Reaction
  • RNA, Messenger
  • RNA, Catalytic
  • Protozoan Proteins
  • Patch-Clamp Techniques
  • Myotonia Congenita
  • Mutation
  • Muscle, Skeletal
 

Citation

APA
Chicago
ICMJE
MLA
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Rogers, C. S., Vanoye, C. G., Sullenger, B. A., & George, A. L. (2002). Functional repair of a mutant chloride channel using a trans-splicing ribozyme. J Clin Invest, 110(12), 1783–1789. https://doi.org/10.1172/JCI16481
Rogers, Christopher S., Carlos G. Vanoye, Bruce A. Sullenger, and Alfred L. George. “Functional repair of a mutant chloride channel using a trans-splicing ribozyme.J Clin Invest 110, no. 12 (December 2002): 1783–89. https://doi.org/10.1172/JCI16481.
Rogers CS, Vanoye CG, Sullenger BA, George AL. Functional repair of a mutant chloride channel using a trans-splicing ribozyme. J Clin Invest. 2002 Dec;110(12):1783–9.
Rogers, Christopher S., et al. “Functional repair of a mutant chloride channel using a trans-splicing ribozyme.J Clin Invest, vol. 110, no. 12, Dec. 2002, pp. 1783–89. Pubmed, doi:10.1172/JCI16481.
Rogers CS, Vanoye CG, Sullenger BA, George AL. Functional repair of a mutant chloride channel using a trans-splicing ribozyme. J Clin Invest. 2002 Dec;110(12):1783–1789.

Published In

J Clin Invest

DOI

ISSN

0021-9738

Publication Date

December 2002

Volume

110

Issue

12

Start / End Page

1783 / 1789

Location

United States

Related Subject Headings

  • Trans-Splicing
  • Tetrahymena
  • Reverse Transcriptase Polymerase Chain Reaction
  • RNA, Messenger
  • RNA, Catalytic
  • Protozoan Proteins
  • Patch-Clamp Techniques
  • Myotonia Congenita
  • Mutation
  • Muscle, Skeletal