Jb. Chen et al., Development of a mouse cell line containing the phi X174 am3 allele as a target for detecting mutation, MUT RES-GTE, 444(2), 1999, pp. 347-353
Citations number
24
Categorie Soggetti
Molecular Biology & Genetics
Journal title
MUTATION RESEARCH-GENETIC TOXICOLOGY AND ENVIRONMENTAL MUTAGENESIS
Transgenic mice containing multiple copies of the Phi X174 am3 allele are b
eing developed as a model for detecting tissue-specific in vivo mutation. I
n order to derive an analogous system for measuring am3 mutation in vitro,
cells were cultured from 15-day-old C57B1/6J mouse embryos that were homozy
gous for the transgene and these cells were transfected with a plasmid expr
essing the SV40 large T-antigen. Two G418-resistant colonies were isolated
from this culture and expanded to continuously proliferating cell lines (PX
-1 and PX-2). Line PX-2 was treated with up to 1.0 mg/ml of N-ethyl-N-nitro
sourea (ENU), assayed for survival by cloning efficiency after overnight cu
lture, and assayed for am3 mutations after 5 days of culture. Survival decr
eased to 31% at the highest dose of ENU, while mutant frequency increased w
ith dose from approximately 2 X 10(-7) in the untreated cells to 13 X 10(-7
) in cultures treated with 0.6 mg/ml of ENU. PX-2 cells also were treated w
ith 0 and 0.6 mg/ml of ENU and mutant frequency assays were performed after
5, 24, 48 and 72 h of growth. The mutant frequency in the treated culture
increased to 20 X 10(-7) at 48 h and remained approximately the same at 72
h. These results indicate that PX-2 cells should be a useful resource for d
eveloping the in vivo am3 mutant assay and for evaluating the sensitivity o
f the am3 allele to various classes of mutagens. (C) 1999 Elsevier Science
B.V. All rights reserved.