1. The lac Operon Laboratory Purpose: Learning to analyze an example of gene expression Example: lac operon Type of control: negative inducible

2. The Laboratory Worksheet asks: How do mutations affect gene expression of the lac operon?

3. Review: Lac Operon:

4. Mutations and gene expressio of lac operon

5. O c mutation: a cis acting mutation

6. I - mutation a trans acting mutation

7. I s mutation a trans acting mutation

8. 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.

9. Try out your ability to reason through the questions.

10. LactoseNo lactoseLactose I+P+O+Z+Y+I+P+O+Z+Y+ - + I-P+O+Z+Y+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 + O + Z + Y + I + P + O c Z + Y - / I + P + O + Z - Y + I + P + O c Z - Y + / I - P + O + Z + Y - I S P + O + Z + Y - / I - P + O + Z - Y + I + P - O c Z + Y + / I + P + O + Z - Y - Β-galactosidasePermease No Lactose 1 2 3 4 5 6 7 8 9 - +

11. The End.

12. Laboratory Exercise

13. Try out your ability to reason through the questions.

14. LactoseNo lactoseLactose I+P+O+Z+Y+I+P+O+Z+Y+ - + I-P+O+Z+Y+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 c Z + Y - / I + P + O + Z - Y + I + P + O c Z - Y + / I - P + O + Z + Y - I S P + O + Z + Y - / I - P + O + Z - Y + I + P - O c Z + Y + / I + P + O + Z - Y - PROTEIN PRODUCT Β-galactosidasePermease No Lactose - + I+P+O+Z-Y-/ I-P+O+Z+Y+

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

16. 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! I s mutation NO LACTOSE BINDING ACTIVITY but has DNA BINDING ACTIVITY. NO TRANSCRIPTION EVEN WITH INDUCER!

17. 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 mutated Inhibits transcription, even with inducer