DNA notes REVISED Edition!


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DNA notes REVISED Edition!

  1. 2. DNA
  2. 3. Griffith’s Experiment <ul><li>Fredrick Griffith </li></ul><ul><li>1928 British scientist </li></ul><ul><li>Wanted to see why people got sick from bacteria (pneumonia) </li></ul><ul><li>Used mice and a strain of disease causing bacteria </li></ul><ul><ul><li>Harmless strain had rough edges when it was grown on a petri dish </li></ul></ul><ul><ul><li>Disease –causing strain had rough edges when grown on a petri dish </li></ul></ul>
  3. 4. <ul><li>S-strain of bacteria killed mouse (smooth) </li></ul><ul><li>R-strain does NOT kill mouse (rough) </li></ul><ul><li>Heat kills the bacteria </li></ul>
  4. 6. What Griffith’s Experiment Proved <ul><ul><li>Griffith’s hypothesis: </li></ul></ul><ul><ul><ul><li>When live, harmless bacteria and heat-killed bacteria are mixed, the heat-killed bacteria passed on disease-causing information to the live cells of the harmless bacteria, causing harmless bacteria to “transform” into bad bacteria </li></ul></ul></ul><ul><li>Transforming Factor </li></ul><ul><ul><li>This factor was probably a gene because he noticed that the offspring inherited the disease as well </li></ul></ul>
  5. 7. Avery’s Experiments: DNA is the Transforming Factor <ul><li>American biologist Oswald Avery, 1944 </li></ul><ul><li>Was transforming factor DNA or Protein? </li></ul><ul><li>Took Griffith’s heat-killed bacteria and good bacteria and made an extract (aka a juice) </li></ul><ul><ul><li>Treated this extract “protein-destroying enzymes” </li></ul></ul><ul><ul><li>Injected mice with the treated extract </li></ul></ul><ul><li>Did the bacteria still function and kill the mice? </li></ul><ul><ul><li>Yes, the bad bacteria still transformed the harmless bacteria…therefore protein did NOT contain the “transforming factor” </li></ul></ul><ul><li>Treated Griffith’s deadly bacteria and harmless bacteria with “DNA-destroying enzymes” </li></ul><ul><li>Did they still function? </li></ul><ul><ul><li>No, the bacteria was not transformed, therefore, DNA had to be the transforming factor </li></ul></ul>
  6. 8. Avery’s Conclusion <ul><li>DNA is the cell’s genetic material </li></ul><ul><li>Scientists were still skeptical </li></ul><ul><li>Protein is made of 20 a.a. and DNA is only 4 nucleotides…didn’t make sense </li></ul><ul><li>They thought….”DNA is too simple!” </li></ul>
  7. 9. Hershey and Chase’s Experiments <ul><li>1952, American biologists Alfred Hershey and Martha Chase </li></ul><ul><li>Which is the hereditary material: Protein or DNA ??? </li></ul><ul><li>Conducted Experiments using viruses </li></ul><ul><li>Viruses </li></ul><ul><ul><li>Package of nucleic acid wrapped in a protein coat </li></ul></ul><ul><ul><li>Not made of cells </li></ul></ul><ul><ul><li>Can only reproduce by infecting living cell with its genetic info </li></ul></ul><ul><ul><li>Genetic info of virus then tells the cell’s organelles to make more viruses </li></ul></ul><ul><ul><li>Bacteriophage (phage)-virus that infects bacteria…literally means “bacteria eater” </li></ul></ul><ul><ul><ul><li>Virus attaches to the surface of bacteria, injects its genetic material into the bacteria, the viral genes cause many more viruses to be made inside the bacteria until the bacteria burst and hundreds of new viruses are released </li></ul></ul></ul>
  8. 12. Hershey and Chase <ul><li>Experiment 1 </li></ul><ul><ul><li>Treated virus with radioactive sulfur-35 isotope (sulfur is in protein but not DNA) </li></ul></ul><ul><ul><li>Sulfur would attach to protein of virus </li></ul></ul><ul><ul><li>If rad. sulfur was found in bacteria, that means it was the protein coat of virus that contained hereditary information </li></ul></ul><ul><li>Experiment 2 </li></ul><ul><ul><li>Treated virus with radioactive phosphorus-32 isotope (phosphorus is in DNA but not protein) </li></ul></ul><ul><ul><li>Phosphorus would attach to DNA of virus </li></ul></ul><ul><ul><li>If rad. phosphorus was found in bacteria, that means it was the protein coat of virus that contained hereditary information </li></ul></ul><ul><li>Used blender to mix up bacteria and viruses </li></ul><ul><li>Experiment 1=radioactive material was found out side the bacteria cells </li></ul><ul><li>Experiment 2=radioactive material was found inside the bacterial cells </li></ul>
  9. 13. Hershey and Chase’s Experiments
  10. 14. Hershey and Chase <ul><li>Conclusion </li></ul><ul><ul><li>The Phage’s (virus’) DNA entered the bacteria during infection but the protein did not </li></ul></ul><ul><ul><li>DNA must carry the genetic information of the virus </li></ul></ul><ul><ul><li>DNA is the hereditary material </li></ul></ul>
  11. 15. DNA Structure <ul><li>Deoxyribonucleic Acid </li></ul><ul><li>DNA is a polymer made up of many monomers called nucleotides </li></ul><ul><li>Nucleotide contains: </li></ul><ul><ul><li>5-carbon sugar called deoxyribose </li></ul></ul><ul><ul><ul><li>RNA contains RIBOSE sugar instead </li></ul></ul></ul><ul><ul><li>Phosphate group </li></ul></ul><ul><ul><li>One Nitrogenous base (there are 4 types) </li></ul></ul>
  12. 16. <ul><li>Sugar and phosphate make up backbone (sides of ladder) </li></ul><ul><li>Nitrogenous bases make up steps of ladder </li></ul><ul><li>Bases are always paired (Chargaff’s rule) </li></ul>
  13. 19. What are these Nitrogenous bases??? <ul><li>Make up the “steps” of the DNA ladder </li></ul><ul><ul><li>One Step= A Purine + A Pyrimidine </li></ul></ul><ul><li>Purines </li></ul><ul><ul><li>Double ring structure </li></ul></ul><ul><ul><li>A denine </li></ul></ul><ul><ul><li>G uanine </li></ul></ul><ul><li>Pyrimidines </li></ul><ul><ul><li>Single-ring structure </li></ul></ul><ul><ul><li>C ytosine </li></ul></ul><ul><ul><li>T hymine (in DNA only) </li></ul></ul><ul><ul><li>U racil (in RNA only) </li></ul></ul>
  14. 21. <ul><li>Adenine (Purine) binds with Thymine (pyrimidine) </li></ul><ul><ul><li>TWO hydrogen bonds </li></ul></ul><ul><li>Cytosine (Purine) binds with Guanine (pyrimidine) </li></ul><ul><ul><li>THREE hydrogen bonds </li></ul></ul>
  15. 22. Chargaff’s Rule <ul><li>Erwin Chargaff, Am. biochemist </li></ul><ul><li>Years before Watson and Crick </li></ul><ul><li>Percentage of guanine and cytosine in DNA sample were about equal…the same with adenine and thymine </li></ul><ul><li>Chargaff’s Rule: A=T and G=C </li></ul><ul><li>Rule for “base-pairing” </li></ul>
  16. 23. Rosalind Franklin and Maurice Wilkins <ul><li>Rosalind Franklin, 1950s, Brit scientist who studied DNA </li></ul><ul><li>Used X-ray diffraction technique to learn about structure of DNA </li></ul><ul><li>She noticed an X-shaped pattern </li></ul><ul><li>This indicated that DNA had a helical structure </li></ul><ul><li>She did not get a chance to publish her research…and her idea was basically stolen  </li></ul>
  17. 24. Watson and Crick <ul><li>23-year old American Biologist James Watson went to work with English Physicist Francis Crick </li></ul><ul><li>Used information from Rosalind Franklin and Maurice Wilkins (1950’s) </li></ul><ul><ul><li>These two scientists used Rosalind Franklin’s x-ray crystallography images to begin to piece together the DNA structure </li></ul></ul><ul><ul><li>Showed that there were 2 strands twisted around each other </li></ul></ul>
  18. 25. Watson and Crick <ul><li>DNA is a double helix </li></ul><ul><li>Created the first accurate model of DNA using wire </li></ul><ul><li>Bases paired up specifically with each other b/c of hydrogen bonding </li></ul><ul><li>Complementary base-pairing </li></ul><ul><ul><li>A purine with a pyrimidine </li></ul></ul><ul><ul><li>A=T </li></ul></ul><ul><ul><li>G=C </li></ul></ul>
  19. 27. DNA Structure <ul><li>Chromatin wrapped around proteins called histones </li></ul><ul><li>Chromatin condenses to form chromosomes during replication </li></ul>
  20. 28. This is what they already knew from the work of many scientists, about the DNA molecule: <ul><li>DNA is made up of subunits which scientists called nucleotides. </li></ul><ul><li>Each nucleotide is made up of a sugar, a phosphate and a base. </li></ul><ul><li>There are 4 different bases in a DNA molecule: adenine (a purine) cytosine (a pyrimidine) guanine (a purine) thymine (a pyrimidine) </li></ul><ul><li>The number of purine bases equals the number of pyrimidine bases </li></ul><ul><li>The number of adenine bases equals the number of thymine bases </li></ul><ul><li>The number of guanine bases equals the number of cytosine bases </li></ul><ul><li>The basic structure of the DNA molecule is helical, with the bases being stacked on top of each other </li></ul>
  21. 29. Central Dogma of Biology <ul><li>DNA  mRNA  protein </li></ul><ul><li>DNA TRANSCRIBES to mRNA </li></ul><ul><ul><li>Process is called transcription </li></ul></ul><ul><li>mRNA TRANSLATES to proteins </li></ul><ul><ul><li>Process is called translation </li></ul></ul><ul><ul><li>mRNA actually makes amino acids, which come together to make proteins </li></ul></ul>
  22. 33. Nucleic Acids <ul><li>DNA </li></ul><ul><ul><li>Double strand </li></ul></ul><ul><ul><li>Deoxyribose sugar </li></ul></ul><ul><ul><li>A=T </li></ul></ul><ul><ul><li>G=C </li></ul></ul><ul><li>RNA </li></ul><ul><ul><li>Single Strand </li></ul></ul><ul><ul><li>Ribose sugar </li></ul></ul><ul><ul><li>A=U </li></ul></ul><ul><ul><li>G=C </li></ul></ul><ul><ul><li>Uracil is the nitrogenous base used instead of THYMINE </li></ul></ul>
  23. 36. <ul><ul><li>DNA parent strand makes 2 daughter strands…one fast, smart daughter strand (leading) and one, slower, no-fast daughter strand (lagging) </li></ul></ul><ul><ul><li>Leading strand (runs 3’ to 5’) </li></ul></ul><ul><ul><li>Lagging strand (runs 5’ to 3’) </li></ul></ul>
  24. 38. How does DNA replicate itself? <ul><li>Template mechanism </li></ul><ul><ul><li>What is a template??? </li></ul></ul><ul><ul><ul><li>PowerPoint presentations…. </li></ul></ul></ul><ul><ul><li>Like the negative of a photograph </li></ul></ul><ul><li>DNA Replication </li></ul><ul><ul><li>Process of copying the DNA molecule </li></ul></ul><ul><ul><ul><li>What phase of the CELL CYCLE? </li></ul></ul></ul><ul><ul><ul><ul><li>S-phase…. </li></ul></ul></ul></ul><ul><ul><li>2 strands of double helix separate (Unzips) </li></ul></ul><ul><ul><li>Each strand acts as a negative for making the new complementary strand </li></ul></ul><ul><ul><li>Nucleotides line up one by one following base pairing rules </li></ul></ul><ul><ul><li>Enzymes (DNA Polymerase and DNA Ligase) link nucleotides together to form 2 new DNA strands called the daughter strands </li></ul></ul>
  25. 39. DNA Replication Simplified <ul><li>Unzip parent DNA </li></ul><ul><li>Add nucleotides to the 2 template strands of DNA </li></ul><ul><ul><li>DNA parent strand makes 2 daughter strands…one fast, smart daughter strand (leading) and one, slower, no-fast daughter strand (lagging) </li></ul></ul><ul><ul><li>Leading strand (runs 3’ to 5’) </li></ul></ul><ul><ul><li>Lagging strand (runs 5’ to 3’) </li></ul></ul><ul><li>Attach fragments on lagging strand </li></ul>
  26. 40. Origins of Replication <ul><li>Specific site on DNA where replication begins </li></ul><ul><ul><li>DNA Helicase: enzyme that binds to origin site and unwinds DNA in both directions </li></ul></ul><ul><li>Copying goes outward in both directions making replication “bubbles” </li></ul><ul><li>Parent strands open up as daughter strands grow on both sides </li></ul><ul><li>Eukaryotic DNA has many origins of replication on a single DNA strand </li></ul><ul><ul><li>Makes copying faster </li></ul></ul><ul><li>Eventually bubbles merge making two new strands </li></ul><ul><ul><li>Each new strand has a part from the original and a part from the new </li></ul></ul><ul><ul><li>Semi-conservative replication </li></ul></ul><ul><ul><ul><li>Means that parent of the original DNA double helix is always kept as part of the new DNA copies </li></ul></ul></ul>
  27. 41. DNA Polymerases <ul><li>Enzymes </li></ul><ul><li>Make covalent bonds between nucleotides of the new strands </li></ul><ul><li>Fast, accurate process </li></ul><ul><ul><li>Error only one in a billion nucleotides </li></ul></ul><ul><li>Brings over nucleotides to unzipped DNA strand and drops them off </li></ul>
  28. 42. More Players in DNA Replication <ul><li>DNA polymerase can only read a strand that is running 3-prime to 5-prime… </li></ul><ul><ul><li>DNA polymerase works non-stop adding nucleotides onto the strand that runs in the 3’ to 5’ direction </li></ul></ul><ul><ul><li>Therefore, Only one strand is made by a smooth, and continuous process… </li></ul></ul><ul><li>The other strand is put together in bits and pieces… </li></ul><ul><ul><li>Each little section of nucleotides is called an “ Okazaki Fragment ” </li></ul></ul><ul><ul><li>These are then “glued” together to make one, continuous strand in the end by another enzyme… DNA Ligase </li></ul></ul>
  29. 43. Important Enzymes <ul><li>DNA Helicase </li></ul><ul><ul><li>unzips </li></ul></ul><ul><li>DNA Polymerase </li></ul><ul><ul><li>Adds nucleotides </li></ul></ul><ul><li>DNA Ligase </li></ul><ul><ul><li>Attaches/glues </li></ul></ul>
  30. 45. Central Dogma of Biology <ul><li>DNA  mRNA  protein </li></ul><ul><li>DNA TRANSCRIBES to mRNA </li></ul><ul><ul><li>Process is called transcription </li></ul></ul><ul><li>mRNA TRANSLATES to proteins </li></ul><ul><ul><li>Process is called translation </li></ul></ul><ul><ul><li>mRNA actually makes amino acids, which come together to make proteins </li></ul></ul>
  31. 46. DNA  mRNA  amino acids/polypeptide chain (Proteins) <ul><li>DNA codes for an RNA strand </li></ul><ul><li>The every 3 bases on the RNA strand code for a specific amino acid </li></ul><ul><ul><li>CODON : three sequential bases that code for a specific a.a. (20 a.a. total) </li></ul></ul><ul><ul><li>Amino acid are strung together to make a protein (primary structure) </li></ul></ul><ul><li>Change DNA will change RNA which will change amino acids, which change protein </li></ul>
  32. 48. Legend: Transcription of DNA to RNA to protein: This dogma forms the backbone of molecular biology and is represented by four major stages. 1. The DNA replicates its information in a process that involves many enzymes: replication. 2. The DNA codes for the production of messenger RNA (mRNA) during transcription. 3. In eukaryotic cells, the mRNA is processed (essentially by splicing) and migrates from the nucleus to the cytoplasm. 4. Messenger RNA carries coded information to ribosomes. The ribosomes &quot;read&quot; this information and use it for protein synthesis. This process is called translation .
  33. 49. Ala: Alanine  Cys: Cysteine  Asp: Aspartic acid  Glu: Glutamic acid Phe: Phenylalanine  Gly: Glycine His: Histidine  Ile: Isoleucine  Lys: Lysine Leu: Leucine  Met: Methionine Asn: Asparagine Pro: Proline Gln: Glutamine Arg: Arginine Ser: Serine Thr: Threonine Val: Valine Trp: Tryptophane Tyr: Tyrosisne
  34. 50. One Gene, One Polypeptide <ul><li>George Beadle and Edward Tatum </li></ul><ul><li>Am. Geneticists </li></ul><ul><li>1940s </li></ul><ul><li>Orange bread mold, Neurospora crassa </li></ul><ul><ul><li>Studied mutant strains of this mold that could not grow </li></ul></ul><ul><ul><li>Each of the strains lacked a specific enzyme needed to produce some molecule the mold needed to grow </li></ul></ul><ul><ul><li>Each strain was defective in a single, specific gene </li></ul></ul><ul><li>“ One-gene, one-enzyme” hypothesis </li></ul><ul><ul><li>The function of an individual gene is to dictate the production of specific enzyme </li></ul></ul><ul><ul><li>Scientists later learned that genes dictate not just enzymes, but a single polypeptide, so the hypothesis became known as the “One Gene, One Polypeptide” hypothesis </li></ul></ul>
  35. 52. DNA  mRNA  Protein <ul><li>Transcription </li></ul><ul><ul><li>Different form of the same message </li></ul></ul><ul><ul><li>DNA makes single stranded RNA (U replaces T) </li></ul></ul><ul><ul><li>RNA leaves nucleus </li></ul></ul><ul><li>Translation </li></ul><ul><ul><li>Translate from nucleic acid language to amino acid language </li></ul></ul><ul><ul><li>Uses codons , 3-base “word” that codes for specific a.a. </li></ul></ul><ul><ul><ul><li>“ code” for an amino acid </li></ul></ul></ul><ul><ul><li>Several codons make a “sentence” that translates to a polypeptide (protein) </li></ul></ul>
  36. 53. The Genetic Code <ul><li>Am. Biochemist Marshall Nirenberg began to crack the genetic code in the 1960s </li></ul><ul><ul><li>Built RNA model with uracil, called poly U, conducted experiments with it and figured out UUU coded for amino acid phenylalanine </li></ul></ul><ul><ul><li>Scientists used his procedures to figure out the other amino acids represented by codons </li></ul></ul><ul><li>Stop codons : UAA, UGA, UAG </li></ul><ul><ul><li>SIGNAL END OF GENETIC MESSAGE </li></ul></ul><ul><li>Start codon: AUG </li></ul><ul><ul><li>SIGNAL TO START TRANSLATING an RNA transcript </li></ul></ul>
  37. 54. Start Codons <ul><li>UAA </li></ul><ul><li>UGA </li></ul><ul><li>UAG </li></ul>Stop Codons <ul><li>AUG </li></ul>
  38. 55. Three Types of RNA <ul><li>mRNA </li></ul><ul><li>tRNA </li></ul><ul><li>rRNA </li></ul>
  39. 56. Three Types of RNA… #1 <ul><li>mRNA (messanger RNA) </li></ul><ul><ul><li>RNA transcribed from DNA template </li></ul></ul><ul><ul><li>RNA polymerase (enzyme) links RNA nucleotides together </li></ul></ul><ul><ul><li>Modified in nucleus before if exits </li></ul></ul><ul><ul><ul><li>RNA splicing: process in which Introns are removed and exons re joined together to make a continuous coding mRNA molecule </li></ul></ul></ul><ul><ul><li>Introns </li></ul></ul><ul><ul><ul><li>Internal non-coding regions of DNA and mRNA </li></ul></ul></ul><ul><ul><ul><li>Space fillers </li></ul></ul></ul><ul><ul><ul><li>They are cut out of mRNA before it is allowed to leave the nucleus </li></ul></ul></ul><ul><ul><ul><li>Process is called RNA splicing (processing) </li></ul></ul></ul><ul><ul><li>Exons </li></ul></ul><ul><ul><ul><li>Coding region of DNA and mRNA that will be translated (Expressed) </li></ul></ul></ul><ul><ul><ul><li>VERY important part of mRNA…it is carrying the message from DNA (def can’t cut this out) </li></ul></ul></ul>
  40. 57. Three Types of RNA…#2 <ul><li>tRNA (transfer RNA) </li></ul><ul><ul><li>The interpreter </li></ul></ul><ul><ul><li>Translate 3-letter base codes into amino acids </li></ul></ul><ul><ul><li>Carries anti-codon on one end (three letters opposite of what is on mRNA) </li></ul></ul><ul><ul><li>Carries amino acid on other end </li></ul></ul><ul><ul><li>Anti-codon recognizes codon and attaches </li></ul></ul>
  41. 58. Three Types of RNA…#3 <ul><li>rRNA (ribosomal RNA) </li></ul><ul><ul><li>Found in ribosome </li></ul></ul><ul><ul><li>Ribosome composed of 2 subunits: </li></ul></ul><ul><ul><ul><li>Small subunit for mRNA to attach </li></ul></ul></ul><ul><ul><ul><li>Large Subunit for two tRNAs to attach </li></ul></ul></ul><ul><ul><ul><ul><li>“ P” site: holds the tRNA carrying the growing polypeptide chain </li></ul></ul></ul></ul><ul><ul><ul><ul><li>“ A” site: holds the tRNA that is carrying the next a.a. to be added to the chain </li></ul></ul></ul></ul><ul><ul><li>When stop codon (UAA, UAG, UGA) is reached, translation ends and polypeptide is released from tRNA by hydrolysis </li></ul></ul>