In vivo trans-splicing of 5' and 3' segments of pre-mRNA directed by corresponding DNA sequences delivered by gene transfer. Academic Article uri icon

Overview

MeSH

  • Animals
  • Cobra Neurotoxin Proteins
  • Genetic Vectors
  • Humans
  • Liver
  • Plasmids
  • Receptors, Nicotinic
  • Reverse Transcriptase Polymerase Chain Reaction
  • Time Factors

MeSH Major

  • Gene Transfer Techniques
  • RNA Precursors
  • Trans-Splicing

abstract

  • We have developed a new paradigm of in vivo gene transfer termed "segmental trans-splicing" (STS), in which individual "donor" and "acceptor" DNA sequences, delivered in vitro or in vivo, generate pre-mRNAs with 5' and 3' splice signals, respectively, and complementary hybridization domains through which the two pre-mRNAs interact, facilitating trans-splicing of the two mRNA fragments. To demonstrate STS, we used alpha-cobratoxin, a neurotoxin that binds irreversibly to postsynaptic nicotinic acetylcholine receptors. Cells or animals receiving both donor and acceptor plasmids, but neither plasmid alone, yielded RT-PCR products with the correct sequence of mature alpha-cobratoxin mRNA, suggesting that trans-splicing had occurred. Mice receiving intravenous administration of > or = 7.5 microg donor + acceptor plasmids, but not either plasmid alone, died within 6 h. These data demonstrate that segmental trans-splicing occurs in vivo. This approach should permit the intracellular assembly of molecules hitherto too large to be accommodated within current gene transfer vectors.

publication date

  • December 2003

has subject area

  • Animals
  • Cobra Neurotoxin Proteins
  • Gene Transfer Techniques
  • Genetic Vectors
  • Humans
  • Liver
  • Plasmids
  • RNA Precursors
  • Receptors, Nicotinic
  • Reverse Transcriptase Polymerase Chain Reaction
  • Time Factors
  • Trans-Splicing

Research

keywords

  • Journal Article

Identity

Language

  • eng

PubMed ID

  • 14664803

Additional Document Info

start page

  • 999

end page

  • 1008

volume

  • 8

number

  • 6