12. Gene Regulation in Prokaryotes

Suppose the lac operon partial diploid cap⁻ I⁺ P⁺ O⁺ Z⁻ Y⁺/ cap⁺ I⁻ P⁺ O⁺ Z⁺ Y⁻ is grown.

Explain how genetic complementation contributes to the growth habit of this strain.

Suppose the lac operon partial diploid cap⁻ I⁺ P⁺ O⁺ Z⁻ Y⁺/ cap⁺ I⁻ P⁺ O⁺ Z⁺ Y⁻ is grown.

Is transcription of -galactosidase and permease inducible, constitutive, or noninducible?

Suppose the lac operon partial diploid cap⁻ I⁺ P⁺ O⁺ Z⁻ Y⁺/ cap⁺ I⁻ P⁺ O⁺ Z⁺ Y⁻ is grown.

Will this partial diploid strain grow on a lactose medium?

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

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.

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 and glucose.

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 -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.

Genotype                  β-Galactosidase Production
                             Inducer Present       Inducer Absent
1. A⁻B⁺C⁺⁻                          +                              +
2. A⁺B⁺C⁻                           +                              +
3.A⁻B⁺C⁺/A⁺B⁺C⁺                +                              +
4. A⁺B⁺C/A⁺B⁺C⁺⁻                +                              –

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 

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⁺ Oᶜ 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⁺ Oᶜ 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⁻ Oᶜ 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⁺