A trial of somatic gene targeting in vivo with an adenovirus vector

Asami Ino, Yasuhiro Naito, Hiroyuki Mizuguchi, Naofumi Handa, Takao Hayakawa, Ichizo Kobayashi

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Background: Gene targeting in vivo provides a potentially powerful method for gene analysis and gene therapy. In order to sensitively detect and accurately measure designed sequence changes, we have used a transgenic mouse system, MutaMouse, which has been developed for detection of mutation in vivo. It carries bacteriophage lambda genome with lacZ+ gene, whose change to lacZ-negative allele is detected after in vitro packaging into bacteriophage particles. We have also demonstrated that gene transfer with a replication-defective adenovirus vector can achieve efficient and accurate gene targeting in vitro. Methods: An 8 kb long DNA corresponding to the bacteriophage lambda transgene with one of two lacZ-negative single-base-pair-substitution mutant allele was inserted into a replication-defective adenovirus vector. This recombinant adenovirus was injected to the transgenic mice via tail-vein. Twenty-four hours later, genomic DNA was extracted from the liver tissue and the lambda::lacZ were recovered by in vitro packaging. The lacZ-negative phage was detected as a plaque former on agar with phenyl-beta-D-galactoside. Results: The mutant frequency of the lacZ-negative recombinant adenovirus injected mice was at the same level with the control mouse (∼1/10000). Our further restriction analysis did not detect any designed recombinant. Conclusions: The frequency of gene targeting in the mouse liver by these recombinant adenoviruses was shown to be less than 1/20000 in our assay. However, these results will aid the development of a sensitive, reliable and PCR-independent assay for gene targeting in vivo mediated by virus vectors and other means.

Original languageEnglish
Article number8
JournalGenetic Vaccines and Therapy
Volume3
DOIs
Publication statusPublished - 2005 Oct 12

ASJC Scopus subject areas

  • Biotechnology
  • Immunology and Allergy
  • Immunology
  • Molecular Medicine

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