Test ames pdf

If large number of colonies on test plate is observed in comparison to control, then such chemical are said to be mutagens. This may be due to spontaneous point mutation on hisidine encoding gene. Ames test Principle Ames test is developed by Bruce N. Ames in s to test for determining if the chemical is mutagens. This test is based on the principle of reverse mutation or back mutation. So, the test is also known as bacterial reverse mutation assay. Test organism: Ames test uses several strains of bacteria Salmonella, E.

Eg A particular strain of Salmonella Typhimurium carry mutation in gene that encodes histidine. So it is an auxotrophic mutant which loss the ability to synthesize histidine an amino acid utilizing the ingredients of culture media.

Those strains are known as His- and require histidine in growth media. Principle Ames test uses several strains of bacteria Salmonella, E. Point mutations are made in the histidine Salmonella typhimurium or the tryptophan Escherichia coli operon, rendering the bacteria incapable of producing the corresponding amino acid. These mutations result in his- or trp- organisms that cannot grow unless histidine or tryptophan is supplied.

This is to say that when a mutagenic event occurs, base substitutions or frameshifts within the gene can cause a reversion to amino acid prototrophy. This is the reverse mutation. These reverted bacteria will then grow in histidine- or tryptophan-deficient media, respectively.

Method I Isolate an auxotrophic strain of Salmonella Typhimurium for histidine. His-ve II Prepare a test suspension of his-ve Salmonella Typhimurium in a plain buffer with test chemical eg. VII After48 hours count the number of colonies in each plate. Result Interpretation The mutagenicity of chemicals is proportional to number of colonies observed. If there is a large number of colonies on the test plate in comparison to control, then such chemical are said to be mutagens.

Very few numbers of colonies can be seen on control plate also. This may be due to spontaneous point mutation on hisidine encoding gene. Uses While Ames test is used to identify the revert mutations which are present in strains, it can also be used to detect the mutagenicity of environmental samples such as drugs, dyes, reagents, cosmetics, waste water, pesticides and other substances which are easily solubilized in a liquid suspension.

Merits Simple, rapid and robust bacterial assay. Ease and low cost of the test make it invaluable for screening substances in our environment for possible carcinogenicity.