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Journal of Biochemistry Advance Access originally published online on March 23, 2007
Journal of Biochemistry 2007 141(5):675-686; doi:10.1093/jb/mvm074
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© 2007 The Japanese Biochemical Society.

Novel Mutation Assay with High Sensitivity based on Direct Measurement of Genomic DNA Alterations: Comparable Results to the Ames Test

Masae Futakami1, Md Salimullah1, Takashi Miura1, Sumio Tokita2 and Koichi Nishigaki1,3,*

1Department of Functional Materials Science; and 2Department of Applied Chemistry, Saitama University, 255 Shimo-Okubo, Sakura-ku, Saitama 338-8570; and 3Rational Evolutionary Design of Advanced Biomolecules, Saitama Small Enterprise Promotion Corporation, SKIP City, 3-12-18 Kamiaoki, Kawaguchi, Saitama 333-0844, Japan

*To whom correspondence should be addressed. Tel/Fax: +81 48-858-3533, E-mail: koichi{at}fms.saitama-u.ac.jp

Received February 19, 2007; Accepted February 21, 2007


  Abstract

Almost all of the methodologies developed to date to assay the potential mutagenicity of chemical substances are based on detection of altered phenotypic traits. The alternative approach of directly screening the whole genome for mutations is not feasible because of the logistics of carrying out mass sequencing of genes. Here we describe a novel and highly sensitive mutation assay, which we term the ‘genome profiling-based mutation assay’ (GPMA) that directly detects mutations generated in genomic DNA. We used GPMA to detect mutations caused by known mutagens such as AF2 and ethidium bromide even at concentrations of 30 ppb. The number of mutations detected was dependent on the number of generations in culture and the concentrations of the mutagens. Almost complete agreement was observed between GPMA and the Ames test in the discrimination of mutagens (63 out of 64). Owing to the high sensitivity of GPMA, the effects of long-term and low-dose exposures and the influence of chemicals of low solubility can also be screened. Thus, genotype-based GPMA can complement the Ames test, which is the standard technology in this field and is based on phenotypic traits.

Key Words: Ames test, genome profiling, high sensitivity, mutagenic reagents, mutation assay

Abbreviations: EthBr, ethidium bromide; GP, genome profiling; GPMA, genome profiling-based mutation assay; µTGGE, micro temperature gradient gel electrophoresis; PaSS, pattern similarity score; spiddos, species identification dots


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