A genetic method to quantitate induced chromosome breaks using a mouse/human monochromosomal hybrid cell line: Identification of potential clastogenic agents

Abstract

A genetic assay to detect the clastogenic potential of environmental agents is described. This assay is based on the cloning efficiency of cells in a medium that permits the growth of cells following loss of a specific chromosome segment resulting from a chromosome break. For this purpose a mouse/human hybrid cell line R12-2 containing a dominantly marked chromosome 5 as the only human component has been constructed. This chromosome 5 carries two dominant selectable markers, Ecogpt and the gene for sensitivity to diphtheria toxin (DT S). Ecogpt codes for the enzyme xanthine-guanine phosphoribosyltransferase which allows selection for cells containing chromosome 5 or the segment carrying Ecogpt as judged by growth in medium supplemented with mycophenolic acid and xanthine (MX medium). Human cells are sensitive to 10 −13 M DT, whereas mouse cells are resistant to 10 −7 M DT and DT s is expressed as a dominant phenotype. Cultivation of R12-2 cells in the medium containing 10 −13 M DT permit the selection of cells that have lost chromosome 5 or the segment carrying DT s. The presence of two selectable markers on the same chromosome permits the identification and quantitation of cells for the selective loss of a specific chromosome segment. Growth of R12-2 cells in MX medium containing 10 −13 M DT therefore, provides a convenient method to determine the frequency of clastogen induced breaks in chromosome 5. The utility of the proposed genetic assay is assessed using X-irradiation as a model clastogen. Our results clearly show a dose related response that is consistent with cytogenetic observations.