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DNA and Its Role in Heredity. 13 DNA and Its Role in Heredity. 13.1 What Is the Evidence that the Gene Is DNA? 13.2 What Is the Structure of DNA? 13.3 How Is DNA Replicated? 13.4 How Are Errors in DNA Repaired? 13.5 How Does the Polymerase Chain Reaction Amplify DNA?.
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13 DNA and Its Role in Heredity 13.1 What Is the Evidence that the Gene Is DNA? 13.2 What Is the Structure of DNA? 13.3 How Is DNA Replicated? 13.4 How Are Errors in DNA Repaired? 13.5 How Does the Polymerase Chain Reaction Amplify DNA?
13 DNA and Its Role in Heredity Lance Armstrong’s cancer was stopped by a drug called cisplatin that forms linkages between DNA strands and prevents replication. Without DNA replication cells can’t divide, and undergo programmed cell death. Opening Question: How does cisplatin work?
13.1 What Is the Evidence that the Gene Is DNA? By the 1920s it was known that chromosomes consisted of DNA and proteins. A new dye that stained DNA provided evidence that DNA is the genetic material. • It was in the right place • It varied among species • It was present in the right amounts
13.1 What Is the Evidence that the Gene Is DNA? Experimental evidence came from work on two strains of Streptococcus pneumoniae. A substance from cells of one strain (even when dead) could produce a heritable change in the other strain.
13.1 What Is the Evidence that the Gene Is DNA? To identify this substance, Oswald Avery treated samples to destroy different molecules. If DNA was destroyed, the transforming activity was lost. There was no loss of activity with destruction of proteins or RNA.
13.1 What Is the Evidence that the Gene Is DNA? Hershey-Chase experiment: used bacteriophage T2 virus to determine whether DNA, or protein, is the genetic material. Part of the virus enters E. coli cells and converts the cell into a virus replication machine.
13.1 What Is the Evidence that the Gene Is DNA? Bacteriophage were grown with either 35S to label the proteins, or with 32P to label the DNA. After infection, bacterial cells and viral remains were separated—the bacteria cells were labeled with 32P, indicating that DNA had entered the cells.
13.1 What Is the Evidence that the Gene Is DNA? Eukaryotic cells can also be transformed (transfection). A genetic marker (a gene that confers an observable phenotype, such as antibiotic resistance) is used to demonstrate transfection. Any cell can be transfected, even an egg cell, resulting in atransgenic organism.
13.2 What Is the Structure of DNA? The structure of DNA was determined using many lines of evidence. One crucial piece came from X-ray diffraction. A purified substance can be made to form crystals. When X-rays are passed through it, position of atoms is inferred from the pattern of diffraction.
Figure 13.6 X-Ray Crystallography Helped Reveal the Structure of DNA
13.2 What Is the Structure of DNA? Rosalind Franklin prepared crystallographs from DNA samples. Her images suggested a double-stranded helix with 10 nucleotides in each full turn. The diameter of 2 nm suggested that the sugar-phosphate backbone of each strand must be on the outside.
13.2 What Is the Structure of DNA? Chemical composition: Biochemists knew that DNA is a polymer of nucleotides. Each nucleotide consists of deoxyribose, a phosphate group, and a nitrogen-containing base.
13.2 What Is the Structure of DNA? The four different nucleotides differed only in the bases: • Purines: adenine (A), guanine (G) • Pyrimidines: cytosine (C), thymine (T)
13.2 What Is the Structure of DNA? Erwin Chargaff noticed that in all DNA, the amount of purines = the amount of pyrimidines. Chargaff’s rule
13.2 What Is the Structure of DNA? Francis Crick and James Watson used model building, plus the physical and chemical evidence to solve the structure of DNA. They published their results in 1953.
13.2 What Is the Structure of DNA? The X-ray diffraction data indicated that the bases are on the inside and the sugar-phosphate groups on the outside of each strand, and that the chains run in opposite directions—antiparallel.
13.2 What Is the Structure of DNA? Antiparallel chains:
13.2 What Is the Structure of DNA? To satisfy Chargaff’s rule, the model paired a purine on one strand with a pyrimidine on the opposite strand, resulting in uniform width.
13.2 What Is the Structure of DNA? Four key features of DNA structure: • It is a double-stranded helix • It is right-handed • It is antiparallel • The outer edges of the bases are exposed in major and minor grooves
13.2 What Is the Structure of DNA? The two chains are held together by: 1. Hydrogen bonding between bases – complementary base pairing: One purine (A or G) with one pyrimidine (T or C)
13.2 What Is the Structure of DNA? 2. Van der Waals forces between adjacent bases on the same strand. When the base rings come near one another, they tend to stack like poker chips.
13.2 What Is the Structure of DNA? Antiparallel strands: direction of strand is determined by the sugar–phosphate bonds.
13.2 What Is the Structure of DNA? Phosphate groups connect to the 3′ C of one sugar, and the 5′ C of the next sugar. Results in one chain with a free 5′ phosphate group—the 5′ end; The other chain has is a free 3′ hydroxyl group—the 3′ end.
13.2 What Is the Structure of DNA? The backbones of the two DNA strands are closer together on one side of the double helix (forming the minor groove) than on the other (forming the major groove). There are four possible configurations of the base pairs in the grooves.
Figure 13.8 Base pairs in DNA Can Interact with Other Molecules
13.2 What Is the Structure of DNA? The outer edges of the base pairs are exposed and accessible for additional hydrogen bonding. The surfaces of the A-T and C-G base pairs are chemically distinct. Binding of proteins to specific base pair sequences is the key to protein-DNA interactions, which are necessary for the replication and expression of DNA.
13.2 What Is the Structure of DNA? The double-helix structure is essential to DNA function: • Stores genetic information: with millions of nucleotides, the base sequences store a huge amount of information • Susceptible to mutations: alterations in base sequences
13.2 What Is the Structure of DNA? • Precisely replicated in cell division by complementary base pairing • Genetic information is expressed as the phenotype—nucleotide sequence determines sequence of amino acids in proteins
13.3 How Is DNA Replicated? The mechanism of DNA replication was confirmed by replicating DNA in a test tube. Ingredients needed: • Deoxyribonucleoside triphosphates dATP, dCTP, dGTP, and dTTP (dNTPs, the monomers of DNA)
13.3 How Is DNA Replicated? • DNA molecules to serve as templates for the sequence of nucleotides • DNA polymerase enzyme • Salts and a pH buffer These experiments confirmed that DNA contains the information needed for its own replication.
13.3 How Is DNA Replicated? Three possible replication patterns: • Semiconservative: Each parent strand is a template; new molecules have one old and one new strand • Conservative: Original molecule serves as a template only • Dispersive: Fragments of DNA are templates, old and new parts are assembled into new molecules
13.3 How Is DNA Replicated? Meselson and Stahl showed that semiconservative replication was the correct model: E. coli cultures were grown with 15N (a heavy, stable isotope that makes DNA more dense), then transferred to a medium with 14N. DNA densities could only be explained by the semiconservative model.
Working with Data 13.1: The Meselson–Stahl Experiment In the Meselson–Stahl experiment, DNA with 14N was separated from DNA with 15N using an ultracentrifuge to create a density gradient of cesium chloride.
Working with Data 13.1: The Meselson–Stahl Experiment DNA bands from successive generations of E. coli after centrifugation. Plots show quantitative analysis of the bands, where height indicates amount of DNA.
