the lac operon laboratory purpose: learning to analyze an example of gene expression example: lac...

The lac Operon Laboratory

Purpose: Learning to analyze an example of gene expression

Example: lac operonType of control: negative inducible

The Laboratory Worksheet asks:

How do mutations affect gene expression of the lac operon?

Review: Lac Operon:

Mutations and gene expressio of lac operon

Oc mutation: a cis acting mutation

I- mutation a trans acting mutation

Is mutation a trans acting mutation

Lab Worksheet: Haploid and Heterozygous Diploid Mutants

Predict if you expect to obtain: transcription/translation of the lacZ and lacY genes

when lactose is absent or present for the E.coli.

Try out your ability to reason through the questions.

Lactose No lactose Lactose

I+P+O+Z+Y+ - +I-P+O+Z+Y+

I+P-O+Z+Y+

I+P+OcZ+Y+

I+P+O+Z-Y-/ I-P+O+Z+Y+

I+P+OcZ+Y-/ I+P+O+Z-Y+

I+P+OcZ-Y+/ I-P+O+Z+Y-

ISP+O+Z+Y-/ I-P+O+Z-Y+

I+P-OcZ+Y+/ I+P+O+Z-Y-

Β-galactosidase Permease

No Lactose

1

2

3

4

5

6

7

8

9

- +

The End.

Laboratory Exercise

Try out your ability to reason through the questions.

Lactose No lactose Lactose

I+P+O+Z+Y+ - +I-P+O+Z+Y+

I+P-O+Z+Y+

I+P+OcZ+Y+

I+P+OcZ+Y-/ I+P+O+Z-Y+

I+P+OcZ-Y+/ I-P+O+Z+Y-

ISP+O+Z+Y-/ I-P+O+Z-Y+

I+P-OcZ+Y+/ I+P+O+Z-Y-

PROTEIN

PRODUCT Β-galactosidase Permease

No Lactose

- +

I+P+O+Z-Y-/ I-P+O+Z+Y+

The next slides are a “key” to the mutations.

Experimental tests on the repressor: I+ cells synthesize full levels of the lac enzymes only in the

presence of an inducer

I- mutations NO DNA BINDING ACTIVITY, TRANSCRIPTION OCCURS EVEN IN ABSENCE OF INDUCER!

Is mutation NO LACTOSE BINDING ACTIVITY but has DNA BINDING ACTIVITY. NO TRANSCRIPTION EVEN WITH INDUCER!

Experimental mutations to the repressor

Lactose binding site

DNA binding site

I - = DNA binding site mutated, prevents binding, allows transcription

I s=inducer binding region mutatedInhibits transcription, even with inducer