15BIS101F2013MutatLect15

Produces gc to ta transversions species

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Unformatted text preview: . Soak 3 small filters disks in "X" and place them in the Soak center of the three histidine minimal agar plates. center Soak three disks in sterile water and place them in Soak the center of the other three plates as negative controls. controls. Incubate plates at 37°C for 20 hours and count the Incubate number of his+ colonies around each disk. number BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez ©2013 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez 61 Ames Test Enzymes in the rat liver are induced by Arochlor or PCB. BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez ©2013 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez 62 Ames Test of Unknown Compound X 109 Cells of Strains “A”, “B” and “C” added to each plate “A”, his- Mutant A CG GC, 10-8 his- Mutant B deletion, 10-10 X X X 950 0 1 H2O H2O H2O 9 his- Mutant C GC AT, 10-9 0 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez ©2013 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez 2 63 Aflotoxin B1 Toxin produced by certain strains of the fungi Aspergillus flavus and A. Toxin Aspergillus parasiticus. A potent mutagen and carcinogen that commonly contaminates parasiticus potent grains and peanuts. Promotes hepatocarcinogenesis in most mammalian species. Produces GC to TA transversions. species. BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez ©2013 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez 64 Assessing Risk of Different Mutagens # of his+ Revertants Aflotoxin B1 (contaminant of (contaminant peanuts) peanuts) Nitrosamines AF-2 (food preservative) (BBQ meats cosmetics) Tris-BP (flame retardant) Lady Clairol (balsam natural brown) Saccharine (artificial sweetener) EDB (fumigant) Microgram of Mutagen Microgram BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez ©2013 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez 65 Conclusion As shown on the previous slide, compound X is a potent mutagen that As promotes CG to GC transversions 100-fold above the negative controls (i.e., above the normal reversion rate for His- mutant Strain A). This (i.e., compound is a candidate for carcinogenicity testing. Some questions for further consideration include: q q Are all genotoxic compounds equally hazardous to our health? Are What precautions should be taken to regulate these compounds in the What environment? environment? To help answer these questions, Dr. Ames developed the concept of To Human Exposure/Rodent Potency Percentage or HERP%. Human HERP% exposure is defined as the annual exposure per 70kgs (154 lb.) of body weight, while rodent potency is defined as the amount of mutagen needed to cause cancer in laboratory rats. For example, HERP% can be use to compare the potential dangers of aflatoxin, a fungal toxin found in peanut butter and ethylene dibromide, a common insect fumigant. peanut Aflatoxin 0.000064mg/70kg X 100 = 2.1% 0.000064mg/70kg 0.003mg/kg 0.003mg/kg Ethylene dibromide Ethylene 150mg/70kg X 100 = 10,000% 150mg/70kg 1.5mg/kg 1.5mg/kg BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez ©2013 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez 66 Summary: Complementation analysis: Determining the number of genes involved in Determining producing a particular phenotype. producing Molecular Basis of Mutation. Single base pair substitutions, insertions, or Single deletions can change the way the triple code of the correct open reading frame (zero reading frame) is read. These forward mutations can lead to loss of function or a forward gene or create a new function. Suppressor Mutations. Mutations can be reversed by a back mutation or by a Mutations suppressor. A suppressor is a second mutation within the gene (intragenic) or in another gene (extragenic) another Spontaneous Mutations. These mutations are rare and occur as a result of errors These in DNA replication, tautomeric shifts in certain bases, and permanent chemical changes in nucleotides. These changes can be caused by chemicals or by physical agents such as ionizing radiation (e.g, X-rays and ultraviolet light). agents Induced Mutations. Chemical and physical agents can be used to induce (i.e., Chemical increase the rate of) mutations by several orders of magnitude. increase Mutations can be caused by slippage in DNA replication. Mutations slippage Ames Test. Using our knowledge of mutations to monitor mutagens in the Using environment. environment. BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez ©2013 BIS101­001, Spring 2013—Genes and Gene Expression R.L. Rodriguez 67...
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This note was uploaded on 02/12/2014 for the course BIS 101 taught by Professor Simonchan during the Spring '08 term at UC Davis.

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