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10.2: Testing for Mutagenic Chemicals in Bacteria and Mice

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  • Ames test

    Figure 10.2.2 Blue Mouse

    Big Blue mice are transgenic for a segment of DNA that contains the DNA of bacteriophage lambda, a virus that infects E. coli, and which serves here as the vector for 3 genetic elements from the lac operon of E. coli:

    • the lacI gene
    • the operator of the operon
    • the beta-galactosidase (lacZ) gene

    The Assay

    The transgenic mice are given repeated doses of the suspected carcinogen for a week or two. If the chemical is mutagenic, it will cause random mutations throughout the genome of each mouse cell. If a mutation occurs in either the lacI gene (which encodes the lac repressor) or the operator,

    the gene (lacZ) for beta-galactosidase will no longer be repressed. To detect this,

    • The DNA is extracted from the tissues of the treated mouse.
    • The vector is isolated and used to make functional bacteriophages.
    • E. coli cells are mixed with the bacteriophage and spread on a solid culture medium.
    • The bacteriophages infect and destroy ("lyze") the E. coli cells.
    • This causes clear circular zones, called plaques, to appear in a "lawn" of bacteria.
    • Before they die, cells that have been infected by bacteriophages carrying a mutated lacI or operator will produce beta-galactosidase.
    • This reacts with a substrate in the culture medium turning it blue.
    • Bacteriophages with unmutated genes produce colorless plaques because no beta-galactosidase is synthesized.
    • Count both colorless and blue plaques.
    • The number of blue plaques divided by the total number of plaques gives the mutation frequency.
    Figure 10.2.4 Blue Plaque courtesy of Stratagene

    This photograph shows one mutant (blue) plaque on a lawn of E. coli containing many non-mutant (clear) plaques.

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