Ch. 12 - Regulation of Gene Expression in Bacteria and Bacteriophage

Sanders - Genetic Analysis: An Integrated Approach 3rd Edition

Sanders3rd EditionGenetic Analysis: An Integrated ApproachISBN: 9780135564172당신이 사용하는 게 아니라요?교과서 변경

Sanders 3rd Edition

Ch. 12 - Regulation of Gene Expression in Bacteria and Bacteriophage

문제 34e

12장, 문제 34e

Northern blot analysis is performed on cellular mRNA isolated from E. coli. The probe used in the northern blot analysis hybridizes to a portion of the lacY sequence. Below is an example of the gel from northern blot analysis for a wild-type lac⁺ bacterial strain. In this gel, lane 1 is from bacteria grown in a medium containing only glucose (minimal medium). Lane 2 is from bacteria in a medium containing only lactose. Following the style of this diagram, draw the gel appearance for northern blots of the bacteria listed below. In each case, lane 1 is for mRNA isolated after growth in a glucose-containing (minimal) medium, and lane 2 is for mRNA isolated after growth in a lactose-only medium.

lac⁻ bacteria with the genotype I⁺ P⁺ O⁺ Z⁻ Y⁺ that has a polar mutation affecting the lacZ gene

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Understand the context of the problem: Northern blot analysis is used to detect specific RNA sequences in a sample. In this case, the probe hybridizes to the lacY mRNA, which encodes the lactose permease protein. The lac operon is regulated by the presence of glucose and lactose in the medium.

Analyze the genotype provided: The bacteria have the genotype I⁺ P⁺ O⁺ Z⁻ Y⁺. This means the lac operon has a functional repressor (I⁺), promoter (P⁺), and operator (O⁺). However, the lacZ gene is nonfunctional (Z⁻), and the lacY gene is functional (Y⁺). The polar mutation in lacZ affects downstream genes, including lacY.

Consider the effect of the polar mutation: A polar mutation in lacZ disrupts transcription of downstream genes in the operon. Since lacY is downstream of lacZ, the polar mutation prevents lacY mRNA production, even though the lacY gene itself is functional.

Determine the expected results for lane 1 (glucose medium): In the presence of glucose, the lac operon is repressed, and no transcription occurs. Therefore, no lacY mRNA is produced, and the northern blot for lane 1 will show no signal.

Determine the expected results for lane 2 (lactose medium): In the presence of lactose, the lac operon would normally be induced. However, due to the polar mutation in lacZ, transcription of lacY is blocked. Thus, no lacY mRNA is produced, and the northern blot for lane 2 will also show no signal.

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Northern Blotting

Northern blotting is a technique used to detect specific RNA sequences in a sample. It involves the separation of RNA by gel electrophoresis, transfer to a membrane, and hybridization with a labeled probe that binds to the target RNA. This method allows researchers to analyze gene expression by visualizing the presence and size of mRNA transcripts.

Lac Operon

The lac operon is a set of genes in E. coli that are involved in the metabolism of lactose. It includes genes such as lacZ, which encodes β-galactosidase, and lacY, which encodes a lactose permease. The operon is regulated by the presence of lactose and glucose, with lactose acting as an inducer that promotes the expression of these genes when glucose is scarce.

Polar Mutation

A polar mutation affects the expression of downstream genes in an operon due to a mutation in an upstream gene. In the context of the lac operon, a polar mutation in lacZ (e.g., lacZ⁻) can prevent the transcription of downstream genes like lacY, leading to a lack of functional lactose permease. This results in altered mRNA levels and protein production, which can be observed in northern blot analysis.

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교과서 질문

Northern blot analysis is performed on cellular mRNA isolated from E. coli. The probe used in the northern blot analysis hybridizes to a portion of the lacY sequence. Below is an example of the gel from northern blot analysis for a wild-type lac⁺ bacterial strain. In this gel, lane 1 is from bacteria grown in a medium containing only glucose (minimal medium). Lane 2 is from bacteria in a medium containing only lactose. Following the style of this diagram, draw the gel appearance for northern blots of the bacteria listed below. In each case, lane 1 is for mRNA isolated after growth in a glucose-containing (minimal) medium, and lane 2 is for mRNA isolated after growth in a lactose-only medium.

lac⁻ bacteria with the genotype I⁺ P⁺ O⁺ Z⁻ Y⁺

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lac⁻ bacteria with the genotype I⁺ P⁺ O⁺ Z⁺ Y⁺ and a mutation that prevents CAP–cAMP binding to the CAP site

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lac⁺ bacteria with the genotype I⁻ P⁺ O^C Z⁺ Y⁺

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lac⁻ bacteria with the genotype I⁺ P⁺ O^C Z⁻ Y⁻

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교과서 질문

lac⁻ bacteria with the genotype I⁺ P⁻ O^C Z⁺ Y⁺

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교과서 질문

A bacterial inducible operon, similar to the lac operon, contains three genes—R, T, and S—that are involved in coordinated regulation of transcription. One of these genes is an operator region, one is a regulatory protein, and the third produces a structural enzyme. In the table below, '+' indicates that the structural enzyme is synthesized and '−' indicates that it is not produced. Use the information provided to determine which gene is the operator, which produces the regulatory protein, and which produces the enzyme.

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