Sensitivity of the ΦX174am3 allele in relation to the endogenousHprt gene for detecting mutation in transgenic mice

Abstract

Transgenic mice have been developed containing multiple, chromosomally integrated copies of the phiX174 am3 allele that serve as reporters for in vivo mutation at a single A:T basepair. In this study, we examined the relative sensitivity of the am3 transgene for detecting the in vivo mutagenicity of N-ethyl-N-nitrosourea (ENU). Three-week-old male phiX174 mice were treated with 0, 40, and 160 mg/kg of ENU. After 1, 3, 6, and 9 weeks, animals were killed, their spleens removed, and isolated splenocytes were used to measure mutant frequencies (MFs) in both the am3 allele and the endogenous Hprt gene. For animals treated with 40 mg/kg of ENU, the Hprt assay detected an average 22-fold increase over background, while the am3 MFs averaged threefold above background. With the 160 mg/kg dose, the Hprt assay detected a 54-fold average increase, while a sixfold average increase above background was found for the transgenic locus. We conclude that the sensitivity of the am3 assay to ENU was compromised by the presence of ex vivo mutations. Adjustment of am3 MFs to exclude these ex vivo mutants could enhance the sensitivity of the assay.