Lecture on DNA

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DNA Structure and Function : 

DNA Structure and Function Chapter 13

Impacts, IssuesHere Kitty, Kitty, Kitty, Kitty, Kitty : 

Impacts, IssuesHere Kitty, Kitty, Kitty, Kitty, Kitty Clones made from adult cells have problems; the cell’s DNA must be reprogrammed to function like the DNA of an egg

13.1 The Hunt for DNA : 

13.1 The Hunt for DNA Investigations that led to our understanding that DNA is the molecule of inheritance reveal how science advances

First Clues to DNA: around the 1860’s Miescher : 

First Clues to DNA: around the 1860’s Miescher Johannes Freidrich Miescher Lymphocytes were difficult to obtain in sufficient enough numbers to study while leucocytes were known to be the one of the main components in pus and could be obtained from bandages at the nearby hospital. The problem was, however, washing the cells off the bandages without damaging them. Miescher devised different salt solutions eventually producing one with sodium sulfate. The cells were filtered. Since centrifuges were not present at this time the cells were allowed to settle at the bottom of a beaker. He then tried to isolate the nuclei free of cytoplasm. He subjected the purified nuclei to an alkaline extraction followed by acidification resulting in a precipitate being formed which Miescher called nuclein (now known as DNA). He found that this contained phosphorus and nitrogen, but not sulfur.

Early and Puzzling Clues : 

Early and Puzzling Clues 1800s: Miescher found nuclein This later turned out to be DNA (deoxyribonucleic acid) in nuclei Miescher died in obscurity, his work was rediscovered later and was a major clue towards ellucidating the structure of DNA

Frederick Griffith: 1920’s : 

Frederick Griffith: 1920’s Griffith's Experiment The famous experiment was done when Griffith was trying to make a vaccine to prevent pneumonia infections in the "Spanish flu" influenza pandemic after World War I, using two strains of the Streptococcus pneumoniae bacterium. The smooth strain (S strain) had a polysaccharide capsule and was virulent when injected, causing pneumonia and killing mice in a day or two. The capsule is a slimy layer on the cells' surface that allows the bacteria to resist the human immune system. The rough strain (R strain) did not cause pneumonia when injected into mice (it was avirulent), since it lacked a capsule. When the virulent S strain was heated to kill it, and then injected into mice, it produced no ill effects. However, when dead S strain mixed with live R strain was injected into the mouse, the R/S mouse died. After isolating bacteria from the blood of the R/S mice, Griffith discovered that the previously avirulent R bacteria had acquired capsules. The bacteria isolated from the mice infected with the mixture of live type II R and heat-killed type III S were now all of the type III S strain, and maintained this phenotype over many generations. Griffith hypothesized that some "transforming principle" from the heat-killed type III S strain converted the type II R strain into the virulent type III S strain. A German bacteriologist, Fred Neufeld, had discovered the pneumococcal types. Until Griffith's experiment, bacteriologists believed they were fixed and unchangeable from generation to generation [3].

Basic Experimental Procedure by Griffith : 

Basic Experimental Procedure by Griffith Early 1900s: Griffith transferred hereditary material from dead cells to live cells Mice injected with live R cells lived Mice injected with live S cells died Mike injected with killed S cells lived Mice injected with killed S cells and live R cells died; live S cells were found in their blood

Griffith’s Experiments : 

Griffith’s Experiments

Slide 9: 

Fig. 13-2, p. 204 R R S A Mice injected with live cells of harmless strain R do not die. Live R cells are in their blood. B Mice injected with live cells of killer strain S die. Live S cells are in their blood. C Mice injected with heat-killed S cells do not die. No live S cells are in their blood. D Mice injected with live R cells plus heat-killed S cells die. Live S cells are in their blood.

Slide 10: 

Fig. 13-2, p. 204 Stepped Art

Slide 11: 

Griffith hypothesized that some "transforming principle" from the heat-killed type III S strain converted the type II R strain into the virulent type III S strain. A German bacteriologist, Fred Neufeld, had discovered the pneumococcal types. Until Griffith's experiment, bacteriologists believed they were fixed and unchangeable from generation to generation [3].

Ahead of his time………… : 

Ahead of his time………… Posthumous Identification of Griffith's Transforming Principle Frederick Griffith in 1936. Griffith was killed at work in his laboratory in 1941, along with longtime friend, bacteriologist William M. Scott, in London as a result of an air raid in the London Blitz. At the time of his death, Griffith was an obscure scientist, whose monumental discovery of pneumococcal transformation was barely known. His Lancet obituary mentions it in passing as part of a single sentence, while his obituary in the British Medical Journal does not mention it at all.[4] It wasn't until 1944 that Griffith's "transforming principle" was identified as DNA by Oswald Theodore Avery, along with coworkers Colin MacLeod and Maclyn McCarty.[5] All modern molecular biology has evolved from this work.

Avery and McCarty Find the Transforming Principle : 

Avery and McCarty Find the Transforming Principle 1940: Avery and McCarty separated deadly S cells (from Griffith’s experiments) into lipid, protein, and nucleic acid components When lipids, proteins, and RNA were destroyed, the remaining substance, DNA, still transformed R cells to S cells Conclusion: DNA is the “transforming principle”

The Hershey-Chase Experiments : 

The Hershey-Chase Experiments

Confirmation of DNA’s Function : 

Confirmation of DNA’s Function 1950s: Hershey and Chase experimented with bacteriophages (viruses that infect bacteria) Protein parts of viruses, labeled with 35S, stayed outside the bacteria DNA of viruses, labeled with 32P, entered the bacteria Conclusion: DNA, not protein, is the material that stores hereditary information

Slide 16: 

Fig. 13-3a, p. 205

Slide 17: 

Fig. 13-3b, p. 205

Slide 18: 

Fig. 13-3b, p. 205 35S remains outside cells Virus particle coat proteins labeled with 35S DNA being injected into bacterium B In one experiment, bacteria were infected with virus particles labeled with a radioisotope of sulfur (35S). The sulfur had labeled only viral proteins. The viruses were dislodged from the bacteria by whirling the mixture in a kitchen blender. Most of the radioactive sulfur was detected in the viruses, not in the bacterial cells. The viruses had not injected protein into the bacteria.

Slide 19: 

Fig. 13-3c, p. 205

Slide 20: 

Fig. 13-3c, p. 205 Virus DNA labeled with 32P 32P remains inside cells Labeled DNA being injected into bacterium C In another experiment, bacteria were infected with virus particles labeled with a radioisotope of phosphorus (32P). The phosphorus had labeled only viral DNA. When the viruses were dislodged from the bacteria, the radioactive phosphorus was detected mainly inside the bacterial cells. The viruses had injected DNA into the cells—evidence that DNA is the genetic material of this virus.

Slide 21: 

Fig. 13-3, p. 205 Stepped Art

13.1 Key ConceptsDiscovery of DNA’s Function : 

13.1 Key ConceptsDiscovery of DNA’s Function The work of many scientists over more than a century led to the discovery that DNA is the molecule that stores hereditary information about traits

Watson and Crick DNA’s Structure : 

Watson and Crick DNA’s Structure Watson and Crick’s discovery of DNA’s structure was based on almost fifty years of research by other scientists

DNA’s Building Blocks : 

DNA’s Building Blocks Nucleotide A nucleic acid monomer consisting of a five-carbon sugar (deoxyribose), three phosphate groups, and one of four nitrogen-containing bases DNA consists of four nucleotide building blocks Two pyrimidines: thymine and cytosine Two purines: adenine and guanine

Four Kinds of Nucleotides in DNA : 

Four Kinds of Nucleotides in DNA

Chargaff’s Rules : 

Chargaff’s Rules Erwin Chargaff and his colleagues noted "regularities" in the base composition of nucleic acids, which they considered "reflected the existence in all DNA preparations of certain structural principles" (Chargaff, 1950, 1951). The amounts of thymine and adenine in DNA are the same, and the amounts of cytosine and guanine are the same: A = T and G = C Shortly thereafter, this "first parity rule" was dramatically confirmed by the Watson-Crick double-helix model (Watson and Crick, 1953). The proportion of adenine and guanine differs among species

Rosalind Franklin : 

Rosalind Franklin Rosalind Franklin’s research in x-ray crystallography revealed the dimensions and shape of the DNA molecule: an alpha helix This was the final piece of information Watson and Crick needed to build their model of DNA

PHOTO 51 by Rosalind Franklin : 

PHOTO 51 by Rosalind Franklin

Watson and Crick’s DNA Model : 

Watson and Crick’s DNA Model A DNA molecule consists of two nucleotide chains (strands), running in opposite directions and coiled into a double helix Base pairs form on the inside of the helix, held together by hydrogen bonds (A-T and G-C)

Watson and Crick’s Model of DNA (1953) : 

Watson and Crick’s Model of DNA (1953)

Slide 31: 

Fig. 13-5b, p. 207 2-nanometer diameter 0.34 nanometer between each base pair 3.4-nanometer length of each full twist of the double helix The numbers indicate the carbon of the ribose sugars (compare Figure 13.4). The 3’ carbon of each sugar is joined by the phosphate group to the 5’ carbon of the next sugar. These links form each strand’s sugar–phosphate backbone. The two sugar–phosphate backbones run in parallel but opposite directions (green arrows). Think of one strand as upside down compared with the other.

Patterns of Base Pairing : 

Patterns of Base Pairing Bases in DNA strands can pair in only one way A always pairs with T; G always pairs with C The sequence of bases is the genetic code Variation in base sequences gives life diversity

Structure of DNA : 

Structure of DNA

Slide 34: 

Fig. 13-5a, p. 207

Animation: DNA close up : 

Animation: DNA close up

13.2 Key ConceptsDiscovery of DNA’s Structure : 

13.2 Key ConceptsDiscovery of DNA’s Structure A DNA molecule consists of two long chains of nucleotides coiled into a double helix Four kinds of nucleotides make up the chains, which are held together along their length by hydrogen bonds

13.3 DNA Replication and Repair : 

13.3 DNA Replication and Repair A cell copies its DNA before mitosis or meiosis I DNA repair mechanisms and proofreading correct most replication errors

Semiconservative DNA Replication : 

Semiconservative DNA Replication Each strand of a DNA double helix is a template for synthesis of a complementary strand of DNA One template builds DNA continuously; the other builds DNA discontinuously, in segments Each new DNA molecule consist of one old strand and one new strand

Enzymes of DNA Replication : 

Enzymes of DNA Replication DNA helicase Breaks hydrogen bonds between DNA strands DNA polymerase Joins free nucleotides into a new strand of DNA DNA ligase Joins DNA segments on discontinuous strand

DNA Replication : 

DNA Replication

Slide 41: 

Fig. 13-4a, p. 206 adenine (A) deoxyadenosine triphosphate, a purine 5’ or PHOSPHATE end of a nucleotide 3’ or sugar end of a nucleotide

Slide 42: 

Fig. 13-4b, p. 206 guanine (G) deoxyguanosine triphosphate, a purine

Slide 43: 

Fig. 13-4c, p. 206 thymine (T) deoxythymidine triphosphate, a pyrimidine

Slide 44: 

Fig. 13-4d, p. 206 cytosine (C) deoxycytidine triphosphate, a pyrimidine

Slide 45: 

Stepped Art Fig. 13-6, p. 208

Animation: DNA replication details : 

Animation: DNA replication details

Semiconservative Replication of DNA : 

Semiconservative Replication of DNA

Discontinuous Synthesis of DNA : 

Discontinuous Synthesis of DNA

Slide 49: 

Fig. 13-8a, p. 209 A Each DNA strand has two ends: one with a 5’ carbon, and one with a 3’ carbon. DNA polymerase can add nucleotides only at the 3’ carbon. In other words, DNA synthesis proceeds only in the 5’ to 3’ direction.

Slide 50: 

Fig. 13-8b, p. 209 The parent DNA double helix unwinds in this direction. Only one new DNA strand is assembled continuously. 5’ The other new DNA strand is assembled in many pieces. 3’ 3’ Gaps are sealed by DNA ligase. 5’ 3’ 3’ 5’ B Because DNA synthesis proceeds only in the 5’ to 3’ direction, only one of the two new DNA strands can be assembled in a single piece. The other new DNA strand forms in short segments, which are called Okazaki fragments after the two scientists who discovered them. DNA ligase joins the fragments into a continuous strand of DNA.

Checking for Mistakes : 

Checking for Mistakes DNA repair mechanisms DNA polymerases proofread DNA sequences during DNA replication and repair damaged DNA When proofreading and repair mechanisms fail, an error becomes a mutation – a permanent change in the DNA sequence

13.3 Key ConceptsHow Cells Duplicate Their DNA : 

13.3 Key ConceptsHow Cells Duplicate Their DNA Before a cell begins mitosis or meiosis, enzymes and other proteins replicate its chromosome(s) Newly forming DNA strands are monitored for errors Uncorrected errors may become mutations

13.4 Using DNA to Duplicate Existing Mammals : 

13.4 Using DNA to Duplicate Existing Mammals Reproductive cloning is a reproductive intervention that results in an exact genetic copy of an adult individual

Cloning : 

Cloning Clones Exact copies of a molecule, cell, or individual Occur in nature by asexual reproduction or embryo splitting (identical twins) Reproductive cloning technologies produce an exact copy (clone) of an individual

Reproductive Cloning Technologies : 

Reproductive Cloning Technologies Somatic cell nuclear transfer (SCNT) Nuclear DNA of an adult is transferred to an enucleated egg Egg cytoplasm reprograms differentiated (adult) DNA to act like undifferentiated (egg) DNA The hybrid cell develops into an embryo that is genetically identical to the donor individual

A Clone Produced by SCNT : 

A Clone Produced by SCNT

Slide 57: 

Fig. 13-10, p. 211

Animation: How Dolly was created : 

Animation: How Dolly was created

Therapeutic Cloning : 

Therapeutic Cloning Therapeutic cloning uses SCNT to produce human embryos for research purposes Researchers harvest undifferentiated (stem) cells from the cloned human embryos

13.4 Key ConceptsCloning Animals : 

13.4 Key ConceptsCloning Animals Knowledge about the structure and function of DNA is the basis of several methods of making clones, which are identical copies of organisms

13.5 Fame and Glory : 

13.5 Fame and Glory In science, as in other professions, public recognition does not always include everyone who contributed to a discovery Rosalind Franklin was first to discover the molecular structure of DNA, but did not share in the Nobel prize which was given to Watson, Crick, and Wilkins

Rosalind Franklin’s X-Ray Diffraction Image : 

Rosalind Franklin’s X-Ray Diffraction Image Franklin died of cancer at age 37, possibly related to extensive exposure to x-rays

13.5 Key ConceptsThe Franklin Footnote : 

13.5 Key ConceptsThe Franklin Footnote Science proceeds as a joint effort; many scientists contributed to the discovery of DNA’s structure

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