Conditional rescue of fvti deficiency using gene targeted insertion of the tta-tetracycline genetic switch

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Mice totally deficient for FactorVII (FVH) survive embryonic development but suffer severe perinatal bleeding and high neonatal mortality. Of the FVII-deficient neonates that were recovered, 70% died within 24 hours of birth. FVH (-/-) mice that survived beyond 24 hours all expired within 24 days, primarily from macroscopically visible intracranial hematomas. To generate adult FVII deficient mice, the tTA-tetracycline (Tc) genetic switch was employed to generate FVII conditional knockouts. A targeting vector was constructed to replace the entire murine FVII gene with a neoR cassette and both elements of the tTA-Tc switch. These included 1) a tTA-IRES-lacZ transcription unit ligated to the FVII promoter and 2) the tTA responsive promoter, pHCMV-1, linked to a murine fVII cDNA. The targeting vector was introduced into embryonic stem cells and G418 resistant ES cells were screened by Southern blot. ES cells in which the endogenous FVII gene was replaced with the tTA-FVII elements by homologous recombination were introduced into embryos by blastocyst injection. Resulting chimeras were bred to generate mice heterozygous for the wt and tTA-FVII alleles (FVI1). Hétérozygotes have been bred to generate mice homozygous for the tTA-FVII allele. Initial results indicate that in the absence of Tc ( when the tTA-FVII gene should be "on") FVIF"-" 7- 1 mice generate low levels of FVII and are rescued from the consequences of FVII deficiency.

Original languageEnglish (US)
Pages (from-to)258a
Issue number11 PART I
StatePublished - Dec 1 2000
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Immunology
  • Hematology
  • Cell Biology

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