1. The Molecular Basis of Inheritance
Chapter 16

2. Unraveling the mysteries of DNA was one of the top discoveries of all time.

3. In April 1953, Watson and Crick shook the scientific world with an elegant double-helical model for the structure of deoxyribonucleic acid.

4. Your genetic inheritance is your DNA which is contained in the 46 chromosomes you got from your parents… and the mitochondrial DNA you got from your momma!

5. This DNA directs an organism’s development of biochemical, anatomical, physiological and (to some extent) behavioral traits.

6. 16-1 The Road to Discovery
It was with the help of many scientists and their studies that lead to determination of the structure and function of DNA.

7. It was Morgan (the fruit fly guy) that showed that genes was located on chromosomes. But remember, chromosomes are DNA wrapped around proteins.

8. So is it the DNA or the proteins that is the genetic material???

9. Most argued that it had to be protein because proteins are more complex material. I mean, DNA is made just of 4 nitrogenous bases…A,T, C, and G.

10. We now know thanks to the…

11. The discovery of the genetic role of DNA began with research by Fredrick Griffith in Griffith worked with two strains (types) of bacteria… one was pathogenic (disease-causing), the other was harmless.

12. The live harmless strain was mixed with a heat-killed pathogenic strain and injected it into a mouse. The mouse died!!! Mouse killer!

13. EXPERIMENT RESULTS Fig. 16-2
Mixture of heat-killed S cells and living R cells
EXPERIMENT
Heat-killed S cells (control)
Living S cells (control)
Living R cells (control)
Living S cells
RESULTS
Figure 16.2 Can a genetic trait be transferred between different bacterial strains?
Mouse dies
Mouse healthy
Mouse healthy
Mouse dies

14. Sorry…all I get from this is that Fred was a mouse killer!!!
YouTube - AP Bio Video: Frederick Griffith

15. So how did this harmless bacteria suddenly become deadly
So how did this harmless bacteria suddenly become deadly??? Griffith called this phenomenon transformation (a change in genotype and phenotype due to the assimilation of external DNA by a cell).

16. What did he say??? Is that English?

17. Griffith said that the “bad” DNA of the pathogenic bacteria, got into the harmless bacteria turning it into a killer bacteria.

18. So how does it prove it was the DNA that transferred into the bacteria
So how does it prove it was the DNA that transferred into the bacteria? Couldn’t it still have been the protein?? Probably not because bacterial DNA is associated with only a small amount of protein.

19. You would think this would be enough but scientists weren’t buying it
You would think this would be enough but scientists weren’t buying it. In 1944, Oswald Avery, Maclyn McCarty and Colin MacLeod announced that the transforming substance was DNA.

20. Scientists were still skeptical, mainly because little was known about DNA.

21. More evidence for DNA as the genetic material came from studies of viruses that infect bacteria (bacteriophages).

22. Phage head Tail sheath Tail fiber DNA 100 nm Bacterial cell Fig. 16-3
Figure 16.3 Viruses infecting a bacterial cell
DNA
100 nm
Bacterial
cell

23. In 1952, Alfred Hershey and Martha Chase performed experiments showing that DNA is the genetic material of a phage known as T2. They designed an experiment showing that only one of the two components of T2 (DNA or protein) enters a bacteria cell during infection.

24. Fig. 16-4-3 EXPERIMENT Empty protein shell
Radioactivity (phage protein) in liquid
Radioactive protein
Phage
Bacterial cell
Batch 1: radioactive sulfur (35S)
DNA
Phage DNA
Centrifuge
Radioactive DNA
Pellet (bacterial cells and contents)
Figure 16.4 Is protein or DNA the genetic material of phage T2?
Batch 2: radioactive phosphorus (32P)
Centrifuge
Radioactivity (phage DNA) in pellet
Pellet

25. What??? Hershey and Chase??? Does this involve chocolate?
YouTube - Biology Project Hilarious

26. It was known that DNA is a polymer of nucleotides, each consisting of a nitrogenous base, a sugar, and a phosphate group.

27. In 1950, Erwin Chargaff reported that DNA composition varies from one species to the next. This evidence of diversity made DNA a more credible candidate for the genetic material.

28. Chargaff’s rules state that in any species there is an equal number of adenine and thymine bases, and an equal number of guanine and cytosine bases. SO???? This proves that A and T pairs up and C and G pairs up.

29. Sugar–phosphate backbone 5 end Sugar (deoxyribose) 3 end
Fig. 16-5
Sugar–phosphate backbone  end
Nitrogenous bases
Thymine (T)
Adenine (A)
Figure 16.5 The structure of a DNA strand
Cytosine (C)
Phosphate
DNA nucleotide
Sugar (deoxyribose)  end
Guanine (G)

30. At this point, most biologists were convinced that DNA was the genetic material, the challenge was to determine how its structure accounts for its role. We all know that structure dictates function!!!!

31. Rosalind Franklin (and her assistant, Maurice Wilkins) used X-ray crystallography to produce a picture of DNA.

32. (b) Franklin’s X-ray diffraction photograph of DNA
Fig. 16-6
Figure 16.6 Rosalind Franklin and her X-ray diffraction photo of DNA
(a) Rosalind Franklin
(b) Franklin’s X-ray diffraction photograph of DNA

33. Franklin’s images of DNA enabled Watson to deduce that DNA was helical
Franklin’s images of DNA enabled Watson to deduce that DNA was helical. Also, Watson could deduce the width of the helix and spacing of the nitrogen bases. The width suggested that DNA was made up of two strands, forming a double helix.

34. (a) Key features of DNA structure (b) Partial chemical structure
Fig. 16-7
5 end
Hydrogen bond
3 end
1 nm
3.4 nm
Figure 16.7 The double helix
For the Cell Biology Video Stick Model of DNA (Deoxyribonucleic Acid), go to Animation and Video Files.
For the Cell Biology Video Surface Model of DNA (Deoxyribonucleic Acid), go to Animation and Video Files.
3 end
0.34 nm
5 end
(a) Key features of DNA structure
(b) Partial chemical structure
(c) Space-filling model

35. Franklin concluded that there were two antiparallel sugar-phosphate backbones, with the nitrogenous bases paired in the molecule’s interior.

36. At first, Watson and Crick thought the bases paired like with like (A with A, T with T, etc) but such pairing did not result in a uniform width. Pairing purines (A and G) with pyrimidines (T and C) resulted in a uniform width consistent with the X-ray.

37. Purine + purine: too wide
Fig. 16-UN1
Purine + purine: too wide
Pyrimidine + pyrimidine: too narrow
Purine + pyrimidine: width consistent with X-ray data

38. The Watson-Crick model explains Chargaff’s rules (in any organism the amount of A=T, and the amount of G=C.

39. Fig. 16-8 Adenine (A) Thymine (T) Guanine (G) Cytosine (C)
Figure 16.8 Base pairing in DNA
Guanine (G)
Cytosine (C)

40. Let’s bring it all together!
YouTube - The Secret of Life -- Discovery of DNA Structure

41. 16-2 DNA Replication
The relationship between structure and function is found in the double helix. The specific base pairing suggested a possible copying mechanism for DNA.

42. Since the two strands of DNA are complementary, each strand acts as a template for building a new strand in replication. To replicate, the DNA parent molecule unwinds, and two new daughter strands are built based on the base-pairing rules.

43. (b) Separation of strands
Fig A T A T A T A T C G C G C G C G T A T A T A T A A T A T A T A T G C G C G C G C
(a) Parent molecule
(b) Separation of strands
(c) “Daughter” DNA molecules, each consisting of one parental strand and one new strand
Figure 16.9 A model for DNA replication: the basic concept

44. Watson and Crick’s semiconservative model of replication predicts that when a double helix replicates, each daughter molecule will have one old strand (conserved from the parent molecule) and one newly made strand.
YouTube - DNA Structure & Testing : What Is DNA Semi-Conservative Replication?

45. There were two other possible ways that DNA could have replicated
There were two other possible ways that DNA could have replicated. One is called the conservative model were the two parent strands rejoin and the daughter DNA contains two newly made strands. Another possibility was the dispersive model where each strand is a mix of old and new.

46. (a) Conservative model
Fig
First replication
Second replication
Parent cell
(a) Conservative model
(b) Semiconserva- tive model
Figure Three alternative models of DNA replication
(c) Dispersive model

47. Two guys, Meselson and Stahl did experiments that confirmed the semiconservative model.
They labeled the nucleotides of the parent strands with a heavy isotope of nitrogen. The new strands would be built with a lighter nitrogen.

48. The first replication produced a band of hybrid DNA which eliminatinated the conservative model. A second replication produced both light and hybrid DNA, eliminating the dispersive model and supporting the semiconservative model.

49. Bacteria cultured in medium containing 15N 2
Fig a
EXPERIMENT 1
Bacteria cultured in medium containing 15N 2
Bacteria transferred to medium containing 14N
RESULTS 3
DNA sample centrifuged after 20 min (after first application) 4
DNA sample centrifuged after 20 min (after second replication)
Less dense
Figure Does DNA replication follow the conservative, semiconservative, or dispersive model?
More dense

50. Semiconservative model
Fig b
CONCLUSION
First replication
Second replication
Conservative model
Semiconservative model
Figure Does DNA replication follow the conservative, semiconservative, or dispersive model?
Dispersive model

51. Could I please see this experiments???
YouTube - Mesel and Stahl Experiment.mp4

52. The copying of DNA is remarkable in its speed and accuracy
The copying of DNA is remarkable in its speed and accuracy. More than a dozen enzymes and other proteins participates in DNA replication.

53. Replication begins at special sites called origins of replication, where the two DNA strands separated, opening up a replication “bubble”. A eukaryotic chromosome may have hundreds or even thousands of origins of replication.

54. Replication proceeds in both directions from each origin of replication until the entire molecule is copied.

55. Double-stranded DNA molecule
Fig b
Origin of replication
Double-stranded DNA molecule
Parental (template) strand
Daughter (new) strand
0.25 µm
Bubble
Replication fork
Figure Origins of replication in E. coli and eukaryotes
Two daughter DNA molecules
(b) Origins of replication in eukaryotes

56. At the end of each replication bubble is a replication fork, a Y-shaped region where new DNA strands are elongating.

57. Helicases are enzymes that untwist the double helix at the replication fork. Single-strand binding proteins binds to and stabilizes the unattached strands until it can be used as a template. Topoisomerase works ahead of the replication fork breaking, swiveling and rejoining DNA strands.

58. Single-strand binding proteins
Fig
Primase
Single-strand binding proteins 3
Topoisomerase 5 3
RNA primer
Figure Some of the proteins involved in the initiation of DNA replication 5 5 3
Helicase

59. At this point, we have untwisted, unzipped and separated our DNA strands. Now it is time to add complimentary nucleotides to the exposed strands. DNA polymerase has this job…unfortunately it cannot initiate synthesis, it needs a primer.

60. DNA polymerase can only add nucleotides to the 3’ end of an exposed strand. So a temporary “plug” of RNA nucleotides are going to act as a primer on the strand that starts at the 5’ end.

61. Primase (an enzyme) has the job of building and laying down the primer
Primase (an enzyme) has the job of building and laying down the primer. The primer is about 5-10 RNA nucleotides long.

62. Now that the primer has been laid down, both exposed strands can now be replicated by adding complimentary nucleotides. DNA polymerase can lay about 500 nucleotides per second in bacteria and 50 per second in human cells.

63. Each nucleotide that is added to a growing DNA strand is a nucleoside triphosphate called dATP… A WHAT??? A nucleotide with 3 phosphates groups. Remember that a normal nucleotide has a sugar, one phosphate group and a nitrogen base.

64. As each nucleoside is added to the growing end of a DNA strand, the last two phosphate groups are hydrolyzed to form what we call a pyrophosphate. The reaction of this hydrolyzing helps attach the nucleotide to the growing strand.

65. Nucleoside triphosphate
Fig
New strand 5 end
Template strand 3 end
5 end
3 end
Sugar A T A T
Base
Phosphate C G C G G C G C
DNA polymerase
3 end A T A
Figure Incorporation of a nucleotide into a DNA strand T
3 end C
Pyrophosphate C
Nucleoside triphosphate
5 end
5 end

67. The antiparallel structure of the double helix (two strands oriented in opposite directions) affects replication. DNA polymerases add nucleotides only to the free 3’ end of a growing strand; therefore, a new DNA strand can elongate only in the 5’ to 3’ direction.

68. What do you mean we can only added nucleotides to the 3’ end????
YouTube - Biology: DNA Replication: A Summary

69. Along one template of DNA, the DNA polymerase synthesizes a leading strand continuously, moving toward the replication fork.

70. Origin of replication 3 5 RNA primer 5 “Sliding clamp” 3 5
Fig b
Origin of replication 3 5
RNA primer 5
“Sliding clamp” 3 5
DNA pol III
Parental DNA 3 5
Figure Synthesis of the leading strand during DNA replication 5 3 5

71. To elongate the other new strand, called the lagging strand, DNA polymerase must work in the direction away from the replication fork. The lagging strand is synthesized as a series of segments called Okazaki fragments, which are joined together by DNA ligase.

72. Overall directions of replication
Fig
Overview
Origin of replication
Leading strand
Lagging strand
Lagging strand 2 1
Leading strand
Overall directions of replication 3 5 5 3
Template strand 3
RNA primer 3 5 1 5
Okazaki fragment 3 5 3 1 5 5 3 3
Figure 16.6 Synthesis of the lagging strand 2 1 5 3 5 3 5 2 1 5 3 3 1 5 2
Overall direction of replication

73. Single-strand binding protein Overall directions of replication
Fig
Overview
Origin of replication
Leading strand
Lagging strand
Leading strand
Lagging strand
Single-strand binding protein
Overall directions of replication
Helicase
Leading strand 5
DNA pol III 3 3
Primer
Primase 5
Parental DNA 3
Figure A summary of bacterial DNA replication
DNA pol III
Lagging strand 5
DNA pol I
DNA ligase 4 3 5 3 2 1 3 5

74. The proteins that participate in DNA replication form a large complex…a “DNA replication machine”…if you will. They just hang around together until it’s time to go to work.

75. Table 16-1

76. Let’s replicate some DNA…shall we?

77. Because it’s so important that the copying of the DNA be perfect, there is methods of proofreading and repairing.

78. DNA polymerase proofreads newly made DNA, replacing any incorrect nucleotides. In mismatched repair, repair enzymes correct errors in base pairings.

79. DNA can be damaged by chemicals, radioactive emissions, X-rays, and UV light. In nucleotide excision repair, a nuclease cuts out and replaces damages stretches of DNA.

80. Fig. 16-18 Nuclease DNA polymerase DNA ligase
Figure Nucleotide excision repair of DNA damage
DNA ligase

81. Every time a DNA molecule is replicated, it gets shorter because the replication process does not provide a way to compete the 5’ ends.

82. Fig. 16-19 Figure 16.19 Shortening of the ends of linear DNA molecules5
Ends of parental DNA strands
Leading strand
Lagging strand 3
Last fragment
Previous fragment
RNA primer
Lagging strand 5 3
Parental strand
Removal of primers and replacement with DNA where a 3 end is available 5 3
Second round of replication
Figure Shortening of the ends of linear DNA molecules 5
New leading strand 3
New lagging strand 5 3
Further rounds of replication
Shorter and shorter daughter molecules

83. Eukaryotic DNA molecules have at their ends a special nucleotide sequences called telomeres.

84. These added ends don’t prevent the shortening of the DNA molecule but they do postpone the erosion of genes near the ends. So after each replications, new telomeres are added.

85. 16-3 The Chromosome
A bacterial chromosome is a double-stranded, circular DNA molecule associated with a small amount of protein.

86. The DNA in bacteria is “supercoiled” and is found in the region of the cell called the nucleoid.

87. Eukaryotic chromosomes have linear DNA molecules associated with a large amount of protein.

88. Chromatin is a complex of DNA and protein and is found in the nucleus of eukaryotic cells. Histones are proteins that are responsible for the first level of DNA packing in chromatin.

89. Chromatin is organized into fibers
Chromatin is organized into fibers. The 10 n-m fiber is made of DNA wound around histones to form nucleosome “beads”. Nucleosomes are strung together like beads on a string.

90. Nucleosomes, or “beads on a string” (10-nm fiber)
Fig a
Nucleosome
(10 nm in diameter)
DNA double helix (2 nm in diameter) H1
Histone tail
Histones
Figure 16.21a Chromatin packing in a eukaryotic chromosome
DNA, the double helix
Histones
Nucleosomes, or “beads on a string” (10-nm fiber)

91. Interactions between nucleosomes cause the thin fiber to coil or fold into the 30-nm fiber.

92. The 30-nm fiber forms loops that attach to proteins
The 30-nm fiber forms loops that attach to proteins. To form the chromatid, the looped domains coil further to the width of 700 nm.

93. Looped domains (300-nm fiber) Metaphase chromosome
Fig b
Chromatid
(700 nm)
30-nm fiber
Loops
Scaffold
300-nm fiber
Figure 16.21b Chromatin packing in a eukaryotic chromosome
Replicated chromosome (1,400 nm)
30-nm fiber
Looped domains (300-nm fiber)
Metaphase chromosome

94. Chromatin is loosely packed in the nucleus during interphase and condenses prior to mitosis.