Suppose that base substitution mutations sufficient to eliminate the function of the operator regions listed below were to occur. For each case, describe how transcription or life cycle would be affected.

lacO mutation in E. coli

Suppose that base substitution mutations sufficient to eliminate the function of the operator regions listed below were to occur. For each case, describe how transcription or life cycle would be affected.

O_R1 mutation in λ phage

Suppose that base substitution mutations sufficient to eliminate the function of the operator regions listed below were to occur. For each case, describe how transcription or life cycle would be affected.

O_R3 mutation in λ phage

Two different mutations affect P_RE. Mutant 1 decreases transcription from the promoter to 10% of normal. Mutant 2 increases transcription from the promoter to 10 times greater than the wild type. How will each mutation affect the determination of the lytic or lysogenic life cycle in mutant λ phage strains? Explain your answers.

How would mutations that inactivate each of the following genes affect the determination of the lytic or lysogenic life cycle in mutated λ phage strains? Explain your answers.

cI

How would mutations that inactivate each of the following genes affect the determination of the lytic or lysogenic life cycle in mutated λ phage strains? Explain your answers.

cII

How would mutations that inactivate each of the following genes affect the determination of the lytic or lysogenic life cycle in mutated λ phage strains? Explain your answers.

cro

How would mutations that inactivate each of the following genes affect the determination of the lytic or lysogenic life cycle in mutated λ phage strains? Explain your answers.

int

How would mutations that inactivate each of the following genes affect the determination of the lytic or lysogenic life cycle in mutated λ phage strains? Explain your answers.

cII and cro

How would mutations that inactivate each of the following genes affect the determination of the lytic or lysogenic life cycle in mutated λ phage strains? Explain your answers.

N

The bacterial insertion sequence IS10 uses antisense RNA to regulate translation of the mRNA that produces the enzyme transposase, which is required for insertion sequence transposition. Transcription of the antisense RNA gene is controlled by P_OUT, which is more than 10 times more efficient at transcription than the P_IN promoter, which controls transposase gene transcription.

If a mutation reduced the transcriptional efficiency of P_OUT so as to be equal to that of P_IN, what is the likely effect on the transposition of IS10?

The bacterial insertion sequence IS10 uses antisense RNA to regulate translation of the mRNA that produces the enzyme transposase, which is required for insertion sequence transposition. Transcription of the antisense RNA gene is controlled by P_OUT, which is more than 10 times more efficient at transcription than the P_IN promoter, which controls transposase gene transcription.

If a mutation of P_IN eliminates its ability to function in transcription, what is the likely effect on the transposition of IS10?

For an E. coli strain with the lac operon genotype I⁺ P⁺ O⁺ Z⁺ Y⁺, identify the level of transcription of the operon genes in each growth medium listed. Specify transcription as 'none,' 'basal,' or 'activated' for each medium, and provide an explanation to justify your answer.

Growth medium contains lactose and glucose.

For an E. coli strain with the lac operon genotype I⁺ P⁺ O⁺ Z⁺ Y⁺, identify the level of transcription of the operon genes in each growth medium listed. Specify transcription as 'none,' 'basal,' or 'activated' for each medium, and provide an explanation to justify your answer.

Growth medium contains glucose but no lactose.

For an E. coli strain with the lac operon genotype I⁺ P⁺ O⁺ Z⁺ Y⁺, identify the level of transcription of the operon genes in each growth medium listed. Specify transcription as 'none,' 'basal,' or 'activated' for each medium, and provide an explanation to justify your answer.

Growth medium contains lactose but no glucose.

How could antisense RNA be used as an antibiotic? What types of genes would you target using this scheme?

The function of tRNA synthetases is to attach amino acids to tRNAs. Suppose the tRNA synthetase responsible for attaching tryptophan to tRNA is mutated in a bacterial strain, with the result that the tRNA synthetase functions at about 15% of the efficiency of the wild-type tRNA synthetase.

How would this mutation affect attenuation of the tryptophan operon? Explain your answer.

The function of tRNA synthetases is to attach amino acids to tRNAs. Suppose the tRNA synthetase responsible for attaching tryptophan to tRNA is mutated in a bacterial strain, with the result that the tRNA synthetase functions at about 15% of the efficiency of the wild-type tRNA synthetase. Would formation of the 3–4 stem-loop structure in mRNA be more frequent or less frequent in the mutant strain than in the wild-type strain? Why?

The following hypothetical genotypes have genes A, B, and C corresponding to lacI, lacO, and lacZ, but not necessarily in that order. Data in the table indicate whether β-galactosidase is produced in the presence and absence of the inducer for each genotype. Use these data to identify the correspondence between A, B, and C and the lacI, lacO, and lacZ genes. Carefully explain your reasoning for identifying each gene.

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^C Z⁺ Y⁺ 

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⁺ 

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^C Z⁺ Y⁺

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^C Z⁺ Y⁺ 

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 

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^C Z⁻ Y⁻ 

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⁺ and a mutation that prevents CAP–cAMP binding to the CAP site 

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.

For the following lac operon partial diploids, determine whether the synthesis of lacZ mRNA is 'constitutive,' 'inducible,' or 'uninducible,' and indicate whether the partial diploid is or (able or not able to utilize lactose). 

The electrophoresis gel shown in part (a) is from a DNase I footprint analysis of an operon transcription control region. DNA sequence analysis of a 35-bp region is shown in part (b). The control region, labeled with ³²P at one end, is shown in a map in part (c). Separate samples of control-region DNA are exposed to DNase I, and the resulting DNase I–digested DNA is run in separate lanes of the electrophoresis gel. Unprotected DNA is in lane 1, DNA protected by repressor protein is in lane 2, and RNA polymerase–protected DNA is in lane 3. The numbers along the electrophoresis gel correspond to the 35-bp sequence labeled on the map in part (c). Use the information provided to solve the following problems.

Determine the DNA sequence of the 35-bp region examined.

The electrophoresis gel shown in part (a) is from a DNase I footprint analysis of an operon transcription control region. DNA sequence analysis of a 35-bp region is shown in part (b). The control region, labeled with ³²P at one end, is shown in a map in part (c). Separate samples of control-region DNA are exposed to DNase I, and the resulting DNase I–digested DNA is run in separate lanes of the electrophoresis gel. Unprotected DNA is in lane 1, DNA protected by repressor protein is in lane 2, and RNA polymerase–protected DNA is in lane 3. The numbers along the electrophoresis gel correspond to the 35-bp sequence labeled on the map in part (c). Use the information provided to solve the following problems.

Locate the regions of the sequence protected by repressor protein and by RNA polymerase.