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Ch. 11 - Gene Mutation, DNA Repair, and Homologous Recombination
Sanders - Genetic Analysis: An Integrated Approach 3rd Edition
Sanders3rd EditionGenetic Analysis: An Integrated ApproachISBN: 9780135564172Not the one you use?Change textbook
All textbooksSanders 3rd EditionCh. 11 - Gene Mutation, DNA Repair, and Homologous RecombinationProblem 33a
Chapter 11, Problem 33a

In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.
Characterize the expected distribution of colony growth on the four plates. Defend your growth prediction for each plate.

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Understand the Ames test setup: The Ames test is used to assess the mutagenic potential of a compound. It employs mutant strains of Salmonella that cannot synthesize histidine (his-). If a compound induces mutations that revert the bacteria to a functional histidine-synthesizing state (his+), colonies will grow on a histidine-free medium.
Analyze the control plates (Plate 1 and Plate 2): Plate 1 contains Strain A (base-substitution mutation) and S9 extract but no EMS. Plate 2 contains Strain B (frameshift mutation) and S9 extract but no EMS. Since no mutagen (EMS) is present, the expected colony growth on these plates should be minimal, representing spontaneous reversion mutations.
Examine Plate 3 (Strain A with EMS and S9 extract): EMS is an alkylating agent that primarily induces base-substitution mutations. Since Strain A has a base-substitution mutation, EMS is likely to cause reversion mutations, allowing the bacteria to grow on the histidine-free medium. Therefore, you would expect more colony growth on Plate 3 compared to Plate 1.
Examine Plate 4 (Strain B with EMS and S9 extract): Strain B has a frameshift mutation, and EMS is less likely to induce frameshift mutations. Thus, the reversion rate for Strain B on Plate 4 is expected to be low, and the colony growth should be similar to or slightly higher than Plate 2 (spontaneous reversion).
Summarize the expected distribution: Minimal growth is expected on Plates 1 and 2 due to spontaneous reversion. Plate 3 should show significant colony growth due to EMS-induced reversion of the base-substitution mutation in Strain A. Plate 4 should show minimal growth, as EMS is less effective at inducing frameshift reversion in Strain B.

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Key Concepts

Here are the essential concepts you must grasp in order to answer the question correctly.

Ames Test

The Ames test is a widely used method for assessing the mutagenic potential of chemical compounds. It employs specific strains of bacteria, typically Salmonella, that are sensitive to mutations. By observing the growth of these bacteria in the presence of a potential mutagen, researchers can infer whether the compound causes genetic changes that lead to increased colony formation.
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Induced Mutations

Types of Mutations

Mutations can be classified into various types, with base-substitution and frameshift mutations being two primary categories. A base-substitution mutation involves the replacement of one nucleotide with another, potentially altering a single amino acid in a protein. In contrast, a frameshift mutation results from the insertion or deletion of nucleotides, shifting the reading frame and often leading to a completely different and nonfunctional protein.
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Mutations and Phenotypes

S9 Extract

S9 extract is a metabolic activation system derived from the liver of rodents, used in mutagenicity testing to simulate the metabolic processes that occur in living organisms. It contains enzymes that can convert pro-mutagens into their active forms, allowing researchers to assess the mutagenic effects of compounds like ethyl methanesulfonate (EMS) under conditions that mimic biological metabolism.
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Traditional vs. Next-Gen
Related Practice
Textbook Question

Alkaptonuria is a human autosomal recessive disorder caused by mutation of the HAO gene that encodes the enzyme homogentisic acid oxidase. A map of the HAO gene region reveals four BamHI restriction sites (B1 to B4) in the wild-type allele and three BamHI restriction sites in the mutant allele. BamHI utilizes the restriction sequence 5′-GGATCC-3′. The BamHI restriction sequence identified as B3 is altered to 5′-GGAACC-3′ in the mutant allele. The mutation results in a Ser-to-Thr missense mutation. Restriction maps of the two alleles are shown below, and the binding sites of two molecular probes (probe A and probe B) are identified.

DNA samples taken from a mother (M), father (F), and two children (C1 and C2) are analyzed by Southern blotting of BamHI-digested DNA. The gel electrophoresis results are illustrated.

Using A to represent the wild-type allele and a for the mutant allele, identify the genotype of each family member. Identify any family member who is alkaptonuric.

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Textbook Question

Alkaptonuria is a human autosomal recessive disorder caused by mutation of the HAO gene that encodes the enzyme homogentisic acid oxidase. A map of the HAO gene region reveals four BamHI restriction sites (B1 to B4) in the wild-type allele and three BamHI restriction sites in the mutant allele. BamHI utilizes the restriction sequence 5′-GGATCC-3′. The BamHI restriction sequence identified as B3 is altered to 5′-GGAACC-3′ in the mutant allele. The mutation results in a Ser-to-Thr missense mutation. Restriction maps of the two alleles are shown below, and the binding sites of two molecular probes (probe A and probe B) are identified.

DNA samples taken from a mother (M), father (F), and two children (C1 and C2) are analyzed by Southern blotting of BamHI-digested DNA. The gel electrophoresis results are illustrated.

In a separate figure, draw the gel electrophoresis band patterns for all the genotypes that could be found in children of this couple.

419
views
Textbook Question

Alkaptonuria is a human autosomal recessive disorder caused by mutation of the HAO gene that encodes the enzyme homogentisic acid oxidase. A map of the HAO gene region reveals four BamHI restriction sites (B1 to B4) in the wild-type allele and three BamHI restriction sites in the mutant allele. BamHI utilizes the restriction sequence 5′-GGATCC-3′. The BamHI restriction sequence identified as B3 is altered to 5′-GGAACC-3′ in the mutant allele. The mutation results in a Ser-to-Thr missense mutation. Restriction maps of the two alleles are shown below, and the binding sites of two molecular probes (probe A and probe B) are identified.

DNA samples taken from a mother (M), father (F), and two children (C1 and C2) are analyzed by Southern blotting of BamHI-digested DNA. The gel electrophoresis results are illustrated.

Explain how the DNA sequence change results in a Ser-to-Thr missense mutation.

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Textbook Question

In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.

What event is being detected by growth of a colony on any of the four plates?

566
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Textbook Question

In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.

Why is the S9 extract added to each of the plates?

538
views
Textbook Question

In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.

Suppose the compound being tested was proflavin instead of EMS. Would this change the Ames test results? Explain why or why not.

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