CH 10: Molecular Biology of the Gene

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CH 10 Molecular Biology of the Gene

• DNA ? RNA ? Protein

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Sections Covered with my titles for each section

• 10.1 DNA as the genetic material
• 10.2/3 Structure of DNA and RNA
• 10.4/5 DNA replication
• 10.6-10.14 Transcription and translation
• 10.15 review
• 10.16 Mutations

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History of DNA

• DNA as the genetic material
• Griffith (1928)
• Found that the genetic component of pathogenic
  bacterial cells was not destroyed when the cells
  were heated
• He did not follow-up on what that component was
  and how/why it survived.
• Griffith Experiment

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DNA as the Genetic Material

• Avery (1944)
• Most believed protein to be genetic material at
  this time.
• Avery found that pathogenic bacterial cells
  treated with protein digesting enzymes could
  still transform harmless bacterial cells.
• Cells treated with a DNA digesting enzyme could
  not.

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DNA as the Genetic Material

• Avery (1944)
• Avery concluded that DNA and not protein must be
  the genetic material.
• Many refused to accept this conclusion.
• Thought his findings only applied to bacteria and
  not eukaryotic cells.

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DNA as the Genetic Material

• Hershey-Chase Experiment (1950)
• Their work confirmed to the scientific community
  that DNA was the genetic material.
• Considered an elegant experiment.
• Very simple and demonstrates a great deal.
• See page 183

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Hershey-Chase Experiment

• They took advantage in a chemical difference
  between DNA and protein
• DNA contains the elements C, H, O, N, P
• Protein contains the elements C, H, O, N, S

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Hershey-Chase Experiment

• Experiment utilized bacteriophages
• Bacteriophages are viruses that infect bacteria.
• Knew that a virus genetic material enters the
  host cell
• as a result the bacterial cell makes more virus
  as directed by the virus genetic material

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Hershey-Chase Experiment

• More on viruses..
• Viruses have two components
• An outer protein coat with nucleic acid inside

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Hershey-Chase Experiment

• The Experiment
• Allowed one sample of viruses to infect bacteria
  grown on a radioactive (RA) sulfur-35 medium
• Viruses made had RA Sulfur-35 in their protein
  coats.

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Hershey-Chase Experiment

• The Experiment
• Allowed another sample of viruses to infect
  bacteria grown on a radioactive (RA)
  phosphorus-32 medium
• Viruses made had RA phosphorus-32 in their DNA.

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Hershey-Chase Experiment

• The two RA viral cultures were isolated and each
  was allowed to infect a new (non RA) bacterial
  culture.
• Expt was done in a liquid medium called the
  supernatant.
• Cultures were gently shaken in a blender to shake
  the virus off of the outside of the bacteria.

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Virus infecting bacterial cell
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Hershey-Chase Experiment

• Each culture was centrifuged to separate the
  liquid medium (supernatant) from the infected
  bacteria.
• The bacteria and the supernatant were checked for
  radioactivity.
• Whatever entered the bacteria is the genetic
  material.

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Hershey-Chase Experiment

• What they found
• Bacteria infected with the virus with a RA S-35
  (protein) coat
• The infected bacteria were NOT RA
• The supernatant was RA
• This is evidence that the protein did not enter
  the bacteria and thus, could not be the genetic
  material.

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Hershey-Chase Experiment

• For the bacteria infected by virus with RA P-32
  in their DNA
• The infected bacteria were RA
• The supernatant was not RA
• This is evidence that the DNA entered the
  bacteria and thus, MUST be the genetic material.
• http//www.accessexcellence.org/RC/VL/GG/hershey.p
  hp

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Structure of DNA

• What was known about DNA
• Chemical components are
• Deoxyribose 5 carbon sugar
• Phosphate groups
• Nitrogenous bases
• Adenine
• Guanine
• Cytosine
• Thymine

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Structure of DNA

• Nitrogenous bases were of 2 types
• Purines have a double-ring structure
• Adenine (A)
• Guanine (G)
• Pyrimidines have a single-ring structure
• Cytosine (C)
• Thymine (T)
• Page 185

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Structure of DNA

• Chargaffs findings (1949)
• Studied DNA from many organisms
• Found that the amount of guanine is always equal
  to the amount cytosine and the amount of adenine
  is equal to the amount of thymine.
• GC
• AT

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Structure of DNA

• X-Ray Crystallography Data on DNA
• Maurice Wilkins and Rosalind Franklin
• Franklins data suggested that DNA was a long
  thin molecule of 2 nm diameter
• Data also indicated a repeating pattern
  consistent with a helix.
• Wilkins shared Franklins data and lab notes with
  Watson and Crick without her permission.

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Rosalind Franklin

• As a scientist Miss Franklin was distinguished by
  extreme clarity and perfection in everything she
  undertook. Her photographs are among the most
  beautiful X-ray photographs of any substance ever
  taken. Their excellence was the fruit of extreme
  care in preparation and mounting of the specimens
  as well as in the taking of the photographs. --
  J. D. Bernal 1958 N

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Franklins X-Ray Data
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Structure of DNA

• "The instant I saw the picture my mouth fell open
  and my pulse began to race.... the black cross of
  reflections which dominated the picture could
  arise only from a helical structure... mere
  inspection of the X-ray picture gave several of
  the vital helical parameters." Watson

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Structure of DNA

• In 1953 Watson, Crick, and Wilkins put the pieces
  together and proposed their famous double helix
  structure for DNA.
• Watson, Crick, and Wilkins were awarded a Nobel
  Prize for deciphering the structure of DNA

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Watson and Crick
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Structure of DNA

• DNA is a double-stranded helix
• Each strand is a long chain of covalently bonded
  nucleotides
• Phosphates can bond to carbon 5 or carbon 3 of
  deoxyribose
• Phoshpates link the sugars to form the backbone
  of the chain
• Bases bond to carbon 1 of deoxyribose
• Page 187

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Structure of DNA

• Each strand has a 5 and a 3 end
• Two DNA strands run in opposite directions
• One runs 5 ? 3 and the other 3? 5

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Structure of DNA

• The two strands are joined by hydrogen bonds
  between the bases
• Two H bonds form between A and T.
• Three H bonds form between G and C.

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C
G
A
T
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Structure DNA
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DNA Replication

• DNA replication DNA synthesis
• Occurs in the nucleus during ___ of the cell
  cycle
• Goal is to make an exact copy of the cells DNA
• Put another way -- goal is to duplicate the
  chromosomes.

Replication
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Semi-Conservative Model

• Each newly made piece of DNA is ½ old DNA and ½
  new DNA (page 188)

Simple animation of replication
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DNA Replication-enzymes needed

• Helicases
• Open the H bonds between the strands
• Stabilizing proteins
• Hold the two strands apart

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DNA Replication enzymes needed

• DNA polymerase III
• Adds nucleotides to the 3 end of DNA
• Saysynthesizes DNA in the 5 ? 3 direction
• It cannot initiate (start) a new DNA strand
• DNA polymerase I
• Removes primer sequences and fills in the gaps
  with DNA
• Other DNA polymerases
• Proofread the DNA and correct mutations

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DNA Replication-enzymes needed

• Primer enzyme not shown in text
• Starts synthesis in the 5 ? 3 direction
• Makes a primer sequence to which DNA polymerase
  III can add DNA
• DNA ligase
• Joins newly made DNA segments after the primer
  sequences have been removed and replaced by DNA
  polymerase I.

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DNA Replication

• Helicases and stabilizing proteins open and
  unwind small sections of DNA and hold the strands
  apart.
• Occurs at specific locations on the DNA called
  origins of replication
• Primer enzymes synthesize primer strands in the
  5 ? 3 direction on each DNA strand.

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DNA Replication

• DNA polymerase III adds DNA to each primer
  sequence in the 5 ? 3 direction.

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DNA Replication

• Proteins open more of the DNA (replication fork
  opens more).
• DNA synthesis continues in the 5? 3 direction
  on one strand. (leading strand)
• Another primer is laid down on the other strand
  and then DNA synthesis continues. (lagging
  strand)

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Primer sequences
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DNA Replication

• Process continues until all of the DNA has been
  replicated.
• Primer sequences are cut out, the gaps filled in
  with DNA
• DNA ligase joins the new DNA sequences.
• http//highered.mcgraw-hill.com/olc/dl/120076/bio2
  3.swfAnimation