Establishment of a Method for Analyzing Translesion DNA Synthesis across a Single Bulky Adduct in Human Cells

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  • Sawai Tomoko
    Environmental Genetics Laboratory, Frontier Science Innovation Center and Department of Biology, Graduate School of Science, Osaka Prefecture University
  • Kawanishi Masanobu
    Environmental Genetics Laboratory, Frontier Science Innovation Center and Department of Biology, Graduate School of Science, Osaka Prefecture University
  • Takamura-Enya Takeji
    Department of Applied Chemistry, Kanagawa Institute of Technology
  • Yagi Takashi
    Environmental Genetics Laboratory, Frontier Science Innovation Center and Department of Biology, Graduate School of Science, Osaka Prefecture University

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  • Establishment of a method for analyzing translesion synthesis across a single bulky adduct in human cells

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Abstract

Translesion DNA synthesis (TLS) is a tolerance pathway of replication block caused by DNA lesions. To measure the efficiency and fidelity of TLS in human cells, we established a shuttle vector assay by modifying of a bacterial TLS assay system. The assay consists of transfection of DNA repair-deficient human cells with a plasmid possessing a single DNA adduct, and transformation of indicator bacteria with plasmids extracted from the cells. We show that plasmid replication was suppressed to 1/9 by a single aminobiphenyl-dG adduct, and the mutant frequency of TLS-operated plasmids was 0.31, of which the major mutation (78%) was G to T transversion. The results demonstrate that this assay is applicable in practice for investigating TLS in human cells.<br>

Journal

  • Genes and Environment

    Genes and Environment 31 (1), 24-30, 2009

    The Japanese Environmental Mutagen Society

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