Detection of integration of plasmid DNA into host genomic DNA following intramuscular injection and electroporation

Z. Wang, P. J. Troilo, X. Wang, T. G. Griffiths, S. J. Pacchione, A. B. Barnum, L. B. Harper, C. J. Pauley, Z. Niu, L. Denisova, T. T. Follmer, G. Rizzuto, G. Ciliberto, E. Fattori, N. L. Monica, S. Manam, B. J. Ledwith

Research output: Contribution to journalArticlepeer-review


Plasmid vectors have been widely used for DNA vaccines and gene therapy. Following intramuscular injection, the plasmid that persists is extrachromosomal and integration into host DNA, if it occurs at all, is negligible. However, new technologies for improving DNA delivery could increase the frequency of integration. In the present study, we tested the effect of electroporation on plasmid uptake and potential integration following intramuscular injection in mice, using a plasmid containing the mouse erythropoietin gene. Electroporation increased plasmid tissue levels by approximately six- to 34-fold. Using a quantitative gel-purification assay for integration, electroporation was found to markedly increase the level of plasmid associated with high-molecular-weight genomic DNA. To confirm integration and identify the insertion sites, we developed a new assay - referred to as repeat-anchored integration capture (RAIC) PCR - that is capable of detecting rare integration events in a complex mixture in vivo. Using this assay, we identified four independent integration events. Sequencing of the insertion sites suggested a random integration process, but with short segments of homology between the vector breakpoint and the insertion site in three of the four cases. This is the first definitive demonstration of integration of plasmid DNA into genomic DNA following injection in vivo.

Original languageEnglish
Pages (from-to)711-721
Number of pages11
JournalGene Therapy
Issue number8
Publication statusPublished - Apr 2004


  • DNA vaccine
  • Electroporation
  • Integration
  • Plasmid

ASJC Scopus subject areas

  • Genetics


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