title: A trial of somatic gene targeting in vivo with an adenovirus vector.
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 (approximately 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.
authors.affiliation: Department of Medical Genome Sciences, Graduate School of Frontier Science, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Tokyo 108-8639, Japan. ino@nibio.go.jp
authors.forename: Asami
authors.initials: A
authors.lastname: Ino
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Naito
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Mizuguchi
authors.forename: Naofumi
authors.initials: N
authors.lastname: Handa
authors.forename: Takao
authors.initials: T
authors.lastname: Hayakawa
authors.forename: Ichizo
authors.initials: I
authors.lastname: Kobayashi