PowerLecture: Chapter 13

1. PowerLecture:Chapter 13 DNA Structure and Function

2. 13.1 The Hunt • Originally believed to be an unknown class of proteins • Thinking was • Heritable traits are diverse • Molecules encoding traits must be diverse • Proteins are made of 20 amino acids and are structurally diverse

3. Miescher Discovered DNA • 1868 • Johann Miescher investigated the chemical composition of the nucleus • Isolated an organic acid that was high in phosphorus • He called it nuclein

4. Griffith Discovers Transformation • 1928 • Attempting to develop a vaccine • Isolated two strains of Streptococcus pneumoniae • Rough strain was harmless • Smooth strain was pathogenic

6. Transformation • What happened in the fourth experiment? • The harmless R cells had been transformed by material from the dead S cells • Descendents of the transformed cells were also pathogenic

7. Avery, McCarty, and MacLeodRepeated Griffith’s Experiment Oswald Avery Maclyn McCarty Colin MacLeod

8. Avery, McCarty, and MacLeodAdded the non-deadly Rough Type of Bacteria to the Heat-Killed Smooth Type To the Heat-Killed Smooth Type, added enzymes that destroyed… Carbohydrates Lipids Proteins RNA DNA

9. S-Type Carbohydrates Destroyed S-Type Lipids Destroyed S-Type Proteins Destroyed S-Type RNA Destroyed S-Type DNA Destroyed Conclusion: DNA was the transforming factor!

10. Oswald & Avery • What is the transforming material? • Cell extracts treated with protein-digesting enzymes could still transform bacteria • Cell extracts treated with DNA-digesting enzymes lost their transforming ability • Concluded that DNA, not protein, transforms bacteria

11. The Hershey-Chase Experiment Protein coat Alfred Hershey & Martha Chase worked with a bacteriophage: A virus that invades bacteria. It consists of a DNA core and a protein coat DNA

12. Protein coats of bacteriophages labeled with Sulfur-35 Phage Hershey and Chase mixed the radioactively-labeled viruses with the bacteria Bacterium Phage The viruses infect the bacterial cells. Bacterium DNA of bacteriophages labeled with Phosphorus-32

13. Protein coats of bacteriophages labeled with Sulfur-35 Separated the viruses from the bacteria by agitating the virus-bacteria mixture in a blender DNA of bacteriophages labeled with Phosphorus-32

14. Protein coats of bacteriophages labeled with Sulfur-35 Centrifuged the mixture so that the bacteria would form a pellet at the bottom of the test tube Measured the radioactivity in the pellet and in the liquid DNA of bacteriophages labeled with Phosphorus-32

15. Hershey & Chase’s Experiments • Created labeled bacteriophages • Radioactive sulfur • Radioactive phosphorus • Allowed labeled viruses to infect bacteria • Asked: Where are the radioactive labels after infection?

16. Hershey and Chase Results 35S remains outside cells virus particle labeled with 35S DNA (blue) being injected into bacterium virus particle labeled with 32P 35P remains inside cells DNA (blue) being injected into bacterium Fig. 13-4ab, p.209

17. Structure of the Hereditary Material • Experiments in the 1950s showed that DNA is the hereditary material • Scientists raced to determine the structure of DNA • 1953 - Watson and Crick proposed that DNA is a double helix Figure 13.6Page 211

18. 13.2 Structure of Nucleotides in DNA • Each nucleotide consists of • Deoxyribose (5-carbon sugar) • Phosphate group • A nitrogen-containing base • Four bases • Adenine, Guanine, Thymine, Cytosine

19. Nucleotide Bases ADENINE (A) GUANINE (G) phosphate group deoxyribose THYMINE (T) CYTOSINE (C)

20. Composition of DNA • Chargaff showed: • Amount of adenine relative to guanine differs among species • Amount of adenine always equals amount of thymine and amount of guanine always equals amount of cytosine A=T and G=C

21. Rosalind Franklin’s Work • Was an expert in X-ray crystallography • Used this technique to examine DNA fibers • Concluded that DNA was some sort of helix

22. Watson-Crick Model • DNA consists of two nucleotide strands • Strands run in opposite directions • Strands are held together by hydrogen bonds between bases • A binds with T and C with G • Molecule is a double helix

23. 13.3 DNA Structure Helps Explain How It Duplicates • DNA is two nucleotide strands held together by hydrogen bonds • Hydrogen bonds between two strands are easily broken • Each single strand then serves as template for new strand

24. How does DNA replicate? Hypotheses: Conservative Semi-Conservative Dispersive

25. Meselson-Stahl Experiment • Bacteria cultured in medium containing a heavy isotope of Nitrogen (15N)‏

26. Meselson-Stahl Experiment • Bacteria transferred to a medium containing elemental Nitrogen (14N)‏

27. Meselson-Stahl Experiment DNA sample centrifuged after First replication

28. Meselson-Stahl Experiment DNA sample centrifuged after Second replication

29. DNA Replication • Each parent strand remains intact • Every DNA molecule is half “old” and half “new” Fig. 13-7, p.212

30. Strand Assembly Why the discontinuous additions? Nucleotides can only be joined to an exposed —OH group that is attached to the 3’ carbon of a growing strand. Energy for strand assembly is provided by removal of two phosphate groups from free nucleotides Fig. 13-8c, p.213

31. Continuous and Discontinuous Assembly • Strands can only be assembled in the 5’ to 3’ direction • continuous on just one parent strand. This is because DNA synthesis occurs only in the 5´ to 3´ direction. • discontinuous: short, separate stretches of nucleotides are added to the template, and then ligase fill in the gaps between them.

32. Base Pairing during Replication Each old strand serves as the template for complementary new strand Fig. 13-8, p. 213

33. Enzymes in Replication • Enzymes unwind the two strands - helicase • DNA polymerase attaches complementary nucleotides • DNA ligase fills in gaps (Okazaki fragments) • Enzymes wind two strands together

34. DNA Repair • Mistakes can occur during replication • DNA polymerase can read correct sequence from complementary strand and, together with DNA ligase, can repair mistakes in incorrect strand

35. 13.4 Cloning • Making a genetically identical copy of an individual • Researchers have been creating clones for decades • These clones were created by embryo splitting

36. Cloning 1 A microneedle 2 The microneedle has emptied the sheep egg of its own nucleus. 3 DNA from a donor cell is about to be deposited in the enucleated egg. 4 An electric spark will stimulate the egg to enter mitotic cell division. the first cloned sheep Fig. 13-9, p.214

37. Dolly: Cloned from an Adult Cell • Showed that differentiated cells could be used to create clones • Sheep udder cell was combined with enucleated egg cell • Dolly is genetically identical to the sheep that donated the udder cell

38. Dolly: Cloned from an Adult Cell Fig. 13-9, p.214

39. Impacts, Issues: Goodbye Dolly • Ian Wilmut was the first to produce a cloned sheep, which he named Dolly • Dolly experienced health problems similar to other mammals cloned from adult DNA

40. Goodbye Dolly Fig. 13-1a, p.206

41. Impacts, Issues: Goodbye Dolly • The risk of defects in clones is huge • Possible benefit – patients in desperate need of organ transplants • Genetically modified cloned animals may produce organs that human donors are less likely to reject • Cloning humans – ethical?

43. Therapeutic Cloning • SCNT – Somatic Cell Nuclear Transfer • Transplant DNA of a somatic cell from the heart, liver, muscles, or nerves into a stem cell (undifferentiated cell)

44. http://www.cyagra.com/gallery/jewel.htm More Clones Cows http://www.popsci.com/scitech/article/2003-05/face-should-we-clone-fading-species

45. Fig. 13-10, p.215