1. DNA: The Carrier of Genetic Information
Chapter 12

2. Learning Objective 1
What evidence was accumulated during the 1940s and early 1950s demonstrating that DNA is the genetic material?

3. The Mystery of Genes
Many early geneticists thought genes were proteins
Proteins are complex and variable
Nucleic acids are simple molecules

4. Evidence for DNA DNA (deoxyribonucleic acid)
Transformation experiments
DNA of one strain of bacteria can transfer genetic characteristics to related bacteria

5. Bacteriophage Experiments
Bacteriophage (virus) infects bacterium
only DNA from virus enters the cell
virus reproduces and forms new viral particles from DNA alone

6. KEY CONCEPTS
Beginning in the 1920s, evidence began to accumulate that DNA is the hereditary material

7. Learning Objective 2
What questions did these classic experiments address?
Griffith’s transformation experiment
Avery’s contribution to Griffith’s work
Hershey–Chase experiments

8. Griffith’s Transformation Experiment
Can a genetic trait be transmitted from one bacterial strain to another?
Answer: Yes

9. Griffith’s Transformation Experiment

10. R cells and heat-killed S cells injected
Experiment 1
Experiment 2
Experiment 3
Experiment 4
R cells injected
S cells injected
Heat-killed
S cells injected
R cells and heat-killed S cells injected
Figure 12.1: Griffith’s transformation experiments.
Griffith was trying to develop a vaccine against pneumonia when he serendipitously discovered the phenomenon of transformation.
Mouse lives
Mouse dies
Mouse lives
Mouse dies
Fig. 12-1, p. 261

11. Animation: Griffith’s Experiment
CLICK TO PLAY

12. Avery’s Experiments
What molecule is responsible for bacterial transformation?
Answer: DNA

13. Hershey–Chase Experiments
Is DNA or protein the genetic material in bacterial viruses (phages)?
Answer: DNA

14. Hershey–Chase Experiments

15. Viruses infect bacteria1
35S
32 P
Bacterial viruses grown in 35S to label protein coat or 32P to label DNA 2
Viruses infect bacteria
Figure 12.2: The Hershey–Chase experiments.
Fig. 12-2, p. 262

16. 3 4 32 P 35S 5 Agitate cells in blender Agitate cells in blender
Separate by centrifugation
Separate by centrifugation
32 P
35S
Figure 12.2: The Hershey–Chase experiments. 5
Bacteria in pellet contain 32P-labeled DNA
35S-labeled protein in supernatant
Fig. 12-2, p. 262

17. Viral reproduction inside bacterial cells from pellet6
Viral reproduction inside bacterial cells from pellet 7
Cell lysis
32P 5
Figure 12.2: The Hershey–Chase experiments. 6 7
Fig. 12-2, p. 262

18. Learning Objective 3
How do nucleotide subunits link to form a single DNA strand?

19. Watson and Crick DNA Model Demonstrated
how information is stored in molecule’s structure
how DNA molecules are templates for their own replication

20. Nucleotides DNA is a polymer of nucleotides
Each nucleotide subunit contains
a nitrogenous base
purines (adenine or guanine)
pyrimidines (thymine or cytosine)
a pentose sugar (deoxyribose)
a phosphate group

21. Forming DNA Chains Backbone Phosphate group attaches to
alternating sugar and phosphate groups
joined by covalent phosphodiester linkages
Phosphate group attaches to
5′ carbon of one deoxyribose
3′ carbon of the next deoxyribose

22. DNA Nucleotides

23. Phosphodiester linkage
Thymine
Adenine
Nucleotide
Phosphate group
Cytosine
Figure 12.3: The nucleotide subunits of DNA.
A single strand of DNA consists of a backbone (superimposed on blue screen) made of phosphate groups alternating with the sugar deoxyribose (green). Phosphodiester linkages (pink) join sugars of adjacent nucleotides. (The nucleotide containing the base adenine is highlighted yellow.) Linked to the 1’ carbon of each sugar is one of four nitrogenous bases (top to bottom): thymine, adenine, cytosine, and guanine. Note the polarity of the polynucleotide chain, with the 5’ end at the top of the figure and the 3’ end at the bottom.
Guanine
Phosphodiester linkage
Deoxyribose (sugar)
Fig. 12-3, p. 264

24. Animation: Subunits of DNA
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25. KEY CONCEPTS
The DNA building blocks consist of four nucleotide subunits: T, C, A, and G

26. Learning Objective 4
How are the two strands of DNA oriented with respect to each other?

27. DNA Molecule
2 polynucleotide chains
associated as double helix

28. DNA Molecule

29. Sugar–phosphate backbone
Minor groove
3.4 nm
Major groove
Figure 12.5: A three-dimensional model of the DNA double helix.
The measurements match those derived from X-ray diffraction images.
0.34 nm
2.0 nm
= hydrogen
= carbon
= oxygen
= atoms in base pairs
= phosphorus
Fig. 12-5, p. 266

30. Double Helix Antiparallel 5′ end 3′ end
chains run in opposite directions
5′ end
phosphate attached to 5′ deoxyribose carbon
3′ end
hydroxyl attached to 3′ deoxyribose carbon

31. KEY CONCEPTS
The DNA molecule consists of two strands that wrap around each other to form a double helix
The order of its building blocks stores genetic information

32. Animation: DNA Close Up
CLICK TO PLAY

33. Learning Objective 5 What are the base-pairing rules for DNA?
How do complementary bases bind to each other?

34. Base Pairs Hydrogen bonding Adenine (A) with thymine (T)
between specific base pairs
binds two chains of helix
Adenine (A) with thymine (T)
forms two hydrogen bonds
Guanine (G) with cytosine (C)
forms three hydrogen bonds

35. Base Pairs and Hydrogen Bonds

36. Fig. 12-6a, p. 267 Figure 12.6: Base pairing and hydrogen bonding.
The two strands of a DNA double helix are hydrogen-bonded between the bases.
Fig. 12-6a, p. 267

37. Adenine Thymine Deoxyribose Deoxyribose Guanine Cytosine Deoxyribose
Figure 12.6: Base pairing and hydrogen bonding.
The two strands of a DNA double helix are hydrogen-bonded between the bases.
Deoxyribose
Deoxyribose
Fig. 12-6b, p. 267

38. Chargaff’s Rules Complementary base pairing
between A and T; G and C
therefore A = T; G = C
If base sequence of 1 strand is known
base sequence of other strand can be predicted

39. KEY CONCEPTS
Nucleotide subunits pair, based on precise pairing rules: T pairs with A, and C pairs with G
Hydrogen bonding between base pairs holds two strands of DNA together

40. Learning Objective 6
What evidence from Meselson and Stahl’s experiment enabled scientists to differentiate between semiconservative replication of DNA and alternative models?

41. Models of DNA Replication

42. (a) Hypothesis 1: Semiconservative replication
Parental DNA
First generation
Second generation
Figure 12.7: Alternative models of DNA replication.
The hypothesized arrangement of old (light blue) and newly synthesized (dark blue) DNA strands after one and two generations, according to (a) the semiconservative model, (b) the conservative model, and (c) the dispersive model.
Fig. 12-7a, p. 268

43. (b) Hypothesis 2: Conservative replication
Parental DNA
First generation
Second generation
Figure 12.7: Alternative models of DNA replication.
The hypothesized arrangement of old (light blue) and newly synthesized (dark blue) DNA strands after one and two generations, according to (a) the semiconservative model, (b) the conservative model, and (c) the dispersive model.
Fig. 12-7b, p. 268

44. (c) Hypothesis 3: Dispersive replication
Parental DNA
First generation
Second generation
Figure 12.7: Alternative models of DNA replication.
The hypothesized arrangement of old (light blue) and newly synthesized (dark blue) DNA strands after one and two generations, according to (a) the semiconservative model, (b) the conservative model, and (c) the dispersive model.
Fig. 12-7c, p. 268

45. Meselson-Stahl Experiment
E. coli
grown in medium containing heavy nitrogen (15N)
incorporated 15N into DNA
Transferred from 15N to 14N medium
after one or two generations, DNA density supported semiconservative replication

46. Meselson-Stahl Experiment

47. Bacteria are grown in 15N (heavy) medium. All DNA is heavy.
Some cells are transferred to 14N (light) medium.
Some cells continue to grow in 14N medium.
Bacteria are grown in 15N (heavy) medium. All DNA is heavy.
First generation
Second generation
High density
Low density
Cesium chloride (CsCl)
DNA
The greater concentration of CsCl at the bottom of the tube is due to sedimentation under centrifigal force.
DNA is mixed with CsCl solution, placed in an ultracentrifuge, and centrifuged at very high speed for about 48 hours.
Figure 12.8: The Meselson–Stahl experiment.
14N (light) DNA
14N – 15N hybrid DNA
15N (heavy) DNA
DNA molecules move to positions where their density equals that of the CsCl solution.
Fig. 12-8a, p. 269

48. One cell generation after transfer to 14N
14N (light) DNA
14N – 15N hybrid DNA
14N – 15N hybrid DNA
15N (heavy) DNA
Figure 12.8: The Meselson–Stahl experiment.
Before transfer to 14N
One cell generation after transfer to 14N
Two cell generations after transfer to 14N
The location of DNA molecules within the centrifuge tube can be determined by UV optics. DNA solutions absorb strongly at 260 nm.
Fig. 12-8b, p. 269

49. Semiconservative Replication
Each daughter double helix consists of
1 original strand from parent molecule
1 new complementary strand

50. Learning Objective 7 How does DNA replicate?
What are some unique features of the process?

51. DNA Replication 2 strands of double helix unwind
each is template for complementary strand
Replication is initiated
DNA primase synthesizes RNA primer
DNA strand grows
DNA polymerase adds nucleotide subunits

52. DNA Replication

53. Base Nucleotide joined to growing chain by DNA polymerase
Figure 12.10: A simplified view of DNA replication.
DNA polymerase adds one nucleotide at a time to the 3’ end of a growing chain.
Phosphates released
Fig , p. 271

54. Other Enzymes DNA helicases Topoisomerases open the double helix
prevent tangling and knotting

55. KEY CONCEPTS
DNA replication results in two identical double-stranded DNA molecules
molecular mechanism passes genetic information from one generation to the next

56. Learning Objective 8
What makes DNA replication (a) bidirectional and (b) continuous in one strand and discontinuous in the other?

57. Bidirectional Replication
Starting at origin of replication
proceeding in both directions
Eukaryotic chromosome
may have multiple origins of replication
may replicate at many points at same time

58. Bidirectional Replication

59. Origin of replication on DNA molecule
DNA polymerase 3’ 3’ 5’ 5’
Figure 12.11: An overview of DNA replication.
Fig a, p. 272

60. Twist introduced into the helix by unwinding
Single-strand binding proteins
RNA primer
DNA polymerase 3’
DNA helicase 3’ 3’ 5’ 3’ 5’
Direction of replication
RNA primer
Figure 12.11: An overview of DNA replication.
Fig b, p. 272

61. 3’ 3’ 5’ 5’ 3’ 3’ 5’ 5’
Figure 12.11: An overview of DNA replication.
Fig c, p. 272

62. DNA Synthesis Always proceeds in 5′ → 3′ direction Leading strand
synthesized continuously
Lagging strand
synthesized discontinuously
forms short Okazaki fragments
DNA primase synthesizes RNA primers
DNA ligase links Okazaki fragments

63. DNA Synthesis

64. Direction of replication 5’ Lagging strand (first Okazaki fragment)3’
Leading strand 5’
DNA helix
RNA primer 3’ 3’
DNA polymerase 5’ 5’
Replication fork 3’
Direction of replication 5’
Lagging strand
(first Okazaki fragment)
Figure 12.12: Leading and lagging DNA strands.
Fig a, p. 273

65. 3’ Leading strand 5’ 3’ RNA primers 3’ 5’ 5’ 3’ 5’ 3’
Figure 12.12: Leading and lagging DNA strands. 5’ 3’
Two Okazaki fragments 5’
Fig b, p. 273

66. Third Okazaki fragment3’ 5’
Leading strand 3’ 3’ 5’ 5’ 3’ 5’
DNA ligase
Third Okazaki fragment
Figure 12.12: Leading and lagging DNA strands.
Lagging strand 3’ 5’
Fig c, p. 273

67. Replication in Bacteria and Eukaryotes

68. Template DNA (light blue)
New DNA (dark blue) 3’ 5’ 5’ 3’
Figure 12.13: Bidirectional DNA replication in bacteria and eukaryotes.
The illustrations do not show leading strands and lagging strands.
Fig a, p. 274

69. Figure 12.13: Bidirectional DNA replication in bacteria and eukaryotes.
The illustrations do not show leading strands and lagging strands.
340 nm
Fig b, p. 274

70. Single replication bubble formed from two merged bubbles 3’ 5’
Single replication bubble formed from two merged bubbles 3’
Replication “bubbles” 3’
Replication fork 5’
Figure 12.13: Bidirectional DNA replication in bacteria and eukaryotes.
The illustrations do not show leading strands and lagging strands.
Fig c, p. 274

71. Animation: Overview of DNA replication and base pairing
CLICK TO PLAY

72. Learning Objective 9
How do enzymes proofread and repair errors in DNA?

73. DNA Polymerases Proofread each new nucleotide
against template nucleotide
Find errors in base pairing
remove incorrect nucleotide
insert correct one

74. DNA Mutation

75. Mutation
Figure 12.9: The perpetuation of a mutation.
The process of DNA replication can stabilize a mutation (bright yellow) so that it is transmitted to future generations.
Fig. 12-9, p. 270

76. Mutation Stepped Art Fig. 12-9, p. 270
Figure 12.9: The perpetuation of a mutation.
The process of DNA replication can stabilize a mutation (bright yellow) so that it is transmitted to future generations.
Stepped Art
Fig. 12-9, p. 270

77. Mismatch Repair
Enzymes recognize incorrectly paired nucleotides and remove them
DNA polymerases fill in missing nucleotides

78. Nucleotide Excision Repair
Repairs DNA lesions
caused by sun or harmful chemicals
3 enzymes
nuclease cuts out damaged DNA
DNA polymerase adds correct nucleotides
DNA ligase closes breaks in sugar–phosphate backbone

79. Nucleotide Excision Repair

80. Nuclease enzyme bound to DNA DNA lesion5’ 3’ 3’ 5’ 5’ 3’ 3’ 5’
DNA polymerase
Figure 12.14: Nucleotide excision repair of damaged DNA.
DNA ligase
New DNA 3’ 5’ 3’ 5’
Fig , p. 275

81. Learning Objective 10 What is a telomere?
What are the possible connections between telomerase and cell aging, and between telomerase and cancer?

82. Telomeres Eukaryotic chromosome ends
noncoding, repetitive DNA sequences
Shorten slightly with each cell cycle
Can be extended by telomerase

83. Replication at Telomeres

84. 5’ 3’ 3’ 5’ DNA replication 5’ 3’ 3’ 5’ RNA primer + RNA primer 5’ 3’
Removal of primer
Figure 12.15: Replication at chromosome ends. 5’ 3’ 3’ 5’ + 5’ 3’ 3’ 5’
Fig a, p. 276

85. 3’ 5’
Figure 12.15: Replication at chromosome ends.
Fig b, p. 276

86. Cell Aging May be caused by absence of telomerase activity
Cells lose ability to divide
after a limited number of cell divisions

87. Cancer Cells Have telomerase Including human cancers
to maintain telomere length and possibly resist apoptosis
Including human cancers
breast, lung, colon, prostate gland, pancreas