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Frequency and spectrum of ENU-induced mutation in the Phi X174 transgene in mouse splenic lymphocytes and their significance to spontaneous transgenic rodent mutation frequencies.

Abstract：

:A perceived disadvantage of transgenic rodent mutation assays is that spontaneous mutant frequencies are high compared to those of endogenous genes and may consequently reduce sensitivity to induced mutation. We have previously argued that unrepaired G:T mismatches from spontaneous deamination of 5-methylcytosine at CpG sites could be converted to apparent in vivo mutations in the bacterial recovery systems because of rapid, random, mismatch repair in Escherichia coli. In this study, we have measured mutation frequencies in spleen of male mice induced by N-ethyl-N-nitrosourea (ENU) using the PhiX174 transgene, which is not subject to mismatch repair in E.coli, using single-burst analysis, a unique method to identify in vivo mutation. In order to compare our results to those using the lacI and cII transgenes, we converted all mutant frequencies to base pair substitution (bps) mutation frequencies per nucleotide based on mutant spectra from this study and published literature. We found this frequency in control spleen to be similar for lacI (3.8 +/- 0.7 x 10(-8)) and PhiX174 (3.1 +/- 1.2 x 10(-8)) at 6 weeks of age. We found a strong age dependence for spontaneous lacI mutation that extrapolated to a value at conception (1.8 +/- 0.9 x 10(-8)) that was not significantly different from the human germ line bps mutation frequency per nucleotide of 1.7 +/- 0.2 x 10(-8). These two transgenes provided similar mutational responses to 40 mg/kg ENU, 7- to 9-fold. In contrast, the cII target gene in the same tissue produces both spontaneous and induced mutation frequencies approximately 10 times higher, for unknown reasons. We conclude that the spontaneous mutant frequencies measured by the lacI and PhiX174 transgenes in this moderately dividing tissue accurately measure in vivo mutation frequencies at early ages. For these two transgenes, seemingly high mutant frequencies may reflect the expected accumulation of somatic mutation with age.

journal_name

Mutagenesis

journal_title

Mutagenesis

authors

Valentine CR,Rainey HF,Farrell JM,Shaddock JG,Dobrovolsky VN,Delongchamp RR

doi

10.1093/mutage/gen026

subject

Has Abstract

pub_date

2008-09-01 00:00:00

pages

383-97

issue

eissn

0267-8357

issn

1464-3804

pii

gen026

journal_volume

pub_type

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Frequency and spectrum of ENU-induced mutation in the Phi X174 transgene in mouse splenic lymphocytes and their significance to spontaneous transgenic rodent mutation frequencies.