title: Site-specific genomic integration produces therapeutic Factor IX levels in mice.
abstract: We used the integrase from phage phiC31 to integrate the human Factor IX (hFIX) gene permanently into specific sites in the mouse genome. A plasmid containing attB and an expression cassette for hFIX was delivered to the livers of mice by using high-pressure tail vein injection. When an integrase expression plasmid was co-injected, hFIX serum levels increased more than tenfold to approximately 4 microg/ml, similar to normal FIX levels, and remained stable throughout the more than eight months of the experiment. hFIX levels persisted after partial hepatectomy, suggesting genomic integration of the vector. Site-specific integration was proven by characterizing and quantifying genomic integration in the liver at the DNA level. Integration was documented at two pseudo-attP sites, native sequences with partial identity to attP, with one site highly predominant. This study demonstrates in vivo gene transfer in an animal by site-specific genomic integration.
authors.affiliation: Department of Genetics, Stanford University School of Medicine, Stanford, CA 94305, USA.
authors.forename: Eric C
authors.initials: EC
authors.lastname: Olivares
authors.forename: Roger P
authors.initials: RP
authors.lastname: Hollis
authors.forename: Thomas W
authors.initials: TW
authors.lastname: Chalberg
authors.forename: Leonard
authors.initials: L
authors.lastname: Meuse
authors.forename: Mark A
authors.initials: MA
authors.lastname: Kay
authors.forename: Michele P
authors.initials: MP
authors.lastname: Calos
mesh_terms: D000818:Animals; D005164:Factor IX / Q000096:biosynthesis* / Q000235:genetics*; D005786:Gene Expression Regulation; D015316:Genetic Therapy / Q000379:methods; D016678:Genome; D007275:Injections, Intravenous; D019426:Integrases / Q000235:genetics / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D016297:Mutagenesis, Site-Directed; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D005164:Factor IX; D019426:Integrases