1. Remember: Final Draft of Posters Due at 10 am tomorrow!
Today:
More Gene Regulation
Intro to Genetic Technology
One last poster team meeting!?
Remember: Final Draft of Posters Due at 10 am tomorrow!

2. Next Up: Regulating at the Level of Transcription!

3. Regulating Transcription
Regulation is almost always based on initiation!

4. Regulating Transcription: Transcription Factors
Transcription factors may be general (required for transcription of all genes) or specific, required for high level expression of particular genes)

5. Regulating Transcription: Enhancers and Activators

6. Evidence for Enhancers
Michael R. Botchan and his colleagues have produced visual evidence of this model of enhancer action. They created an artificial DNA molecule and observed how it interacted with enhancers using an electron microscope

7. Regulating Transcription
Some specific transcription factors function as repressors!
Common Mechanisms:
Block binding of activators
Bind to their own control elements
Recruit histone deacetylases

8. Combinatorial Control
Numbers and diversity of control elements suggest that a particular combination of control elements regulates transcription.

9. Gene Clusters for Coordinated Control
Eukaryotic genes may also be packaged together for regulation. But each has its own promoter and is individually transcribed.
More often, genes in a pathway are scatted on multiple chromosomes, but respond to the same combination of control elements.

10. Post-Transcriptional Regulation

11. Post-Transcription: RNA Processing
(This helps explain our embarrassingly small number of genes!)

12. Post-Transcription: mRNA Degradation
Shortening the poly-A tail can trigger removal of the 5’ cap, followed by breakdown of the mRNA
Hypothesis, TTP promotes mRNA instability.
Prediction: In TTP knocked out, mRNA will…
Figure: Effect of TTP deficiency on the stability of TNF-a mRNA in bone-marrow derived macrophages
J. Blackshear, Biochem. Soc. Trans.. (2001) 30, (945–952)

13. Post-Transcription: RNA Interference (RNAi)
Experimental Observation: Injecting dsRNA into a cell can silence the corresponding gene!

14. Post-Transcription: Initiation of Translation
Regulatory proteins can block attachment of ribosomes
Lack of poly-A tails blocks translation
Block may be global (all translation; i.e. egg cells, dormant plants)

15. Post-Transcription: Protein Processing and Degradation
Many proteins must be cleaved or modified to become active.
Proteins may also be labeled for destruction by attaching a small protein, ubiquitin, that attracts proteosomes

16. Genome Regulation and Cancer
Viruses may carry oncogenes (genes causing cancer)
Oncogenes are similar to proto-oncogenes (genes involved in normal cell division) in our genome

17. A Multistep Model for Cancer

18. Next Up: DNA Technology
What’s interesting, exciting, frightening?
Capture your thoughts: Question 1

19. DNA Tools: Restriction Enzymes
What do Restriction Enzymes do?
Where do they cut (and why)??

20. DNA Tools: Restriction Enzymes

21. DNA Tools: Gel Electorphoresis
You Try! Question 2

22. DNA Tools: Southern Blots
You Try! Question 3

23. DNA Tools: Western and Northern Blots
What can a Northern or Western Blot tell us that a Southern can’t??
You Try: Question 4

24. DNA Tools: Fingerprints
DNA Fingerprints can be made from highly variable regions (polymorphisms).
Two are commonly used: Single Nucleotide Polymorphisms (SNPs) and Short Tandem Repeats (STRs)

25. DNA Tools: Fingerprints
You Try! Question 5

26. Building Recombinant DNA

27. DNA Tools: Vectors Good Vectors: Replicate Independently
Provide Recognition Sequences for Restriction Enzymes
Include a Reporter Gene
Are Small

28. Where do you get DNA to Clone??
DNA (Gene) Libraries?
Are all these clones useful to you?

29. How Could You Get a Library of Expressed, Edited Genes?!?

30. DNA Tools: Knock-Outs

31. DNA Tools: Antisense RNA

32. DNA Tools: Micro-arrays
Let’s watch!

33. Applications: Medically Useful Proteins

34. Applications: Medically Useful Proteins

35. Applications: Pharming
Webster and Pete;
Photo BBC

36. Applications: Agriculture

37. Genetic Technology: DNA Sequencing
Why would we want to sequence DNA?? What enzymes/systems might be helpful?

38. DNA Sequencing, Step 1

39. DNA Sequencing, Step 2

40. DNA Sequencing, Step 2
Normal nucleotide
Dideoxy-nucleotide

41. DNA Sequencing, Step 3

42. Automated DNA Sequencing

43. DNA Sequencing: The Results

44. Applications: Creating Life?!
Let’s Watch!
Capture your thoughts! Question 6