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DNA

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techniques: used a red-dye that binds specifically to DNA ... avirulent form: R form. Discovery of DNA as genetic material. S form: ... – PowerPoint PPT presentation

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Title: DNA


1
DNA
  • the discovery
  • DNA structure
  • DNA replication

2
Discovery of DNA as genetic material
  • 1920s scientists new that chromosomes were made
    up of DNA and protein
  • techniques used a red-dye that binds
    specifically to DNA
  • found that DNA was stored in the nucleus
  • amount of DNA varied by species
  • gametes contained ½ the amount of DNA as somatic
    cells

3
Discovery of DNA as genetic material
  • Still begged the question
  • ? What is the hereditary material the DNA or
    the protein?
  • Two sets of experiments answered this question
    one involving bacteria the other involving
    viruses

4
Discovery of DNA as genetic material
  • Frederick Griffith (1920s) an English physician
  • worked with Streptococcus pneumonia, a bacteria
    that causes pneumonia in humans
  • two forms of the bacteria
  • virulent form S form
  • avirulent form R form

5
Discovery of DNA as genetic material
  • S form
  • contains a polysaccharide capsule
  • gives bacterial colonies a smooth appearance
  • capsule protects the bacterial cell from a
    persons immune system ? virulence factor
  • R form
  • does not contain a polysaccharide capsule
  • has a rough appearance
  • susceptible to attack by the immune system

6
Discovery of DNA as genetic material
  • Griffiths purpose was to develop a vaccine
    against the Streptococcus pneumoniae
  • he inoculated some mice with heat-killed S strain
    ? no infection
  • he infected some mice with a combination of
    heat-killed S strain and R strain ? mice died of
    pneumonia
  • blood of mice full of virulent, S strain bacteria

7
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8
Discovery of DNA as genetic material
  • Griffiths conclusion
  • Some of the R strain bacteria had become
    transformed by the heat-killed S strain into
    virulent S strain bacteria.

9
Discovery of DNA as genetic material
  • Griffiths conclusion did not answer the
    questions of what was genetic material
  • it just established the notion that some
    substance existed the chemical transforming
    principle

10
Discovery of DNA as genetic material
  • Oswald Avery
  • treated samples known to contain pneumoccal
    transforming principle in a variety of ways to
    destroy different types of molecules (proteins,
    nucleic acids, carbohydrates, and lipids)
  • Results If DNA in a sample was destroyed,
    transforming activity was lost but there was no
    loss of activity if one of the other organic
    compounds was destroyed.

11
Discovery of DNA as genetic material
  • Avery and his colleagues published their findings
    in 1944, but results were not seriously
    considered by the scientific community
  • most scientists did not believe DNA was
    chemically complex enough to be genetic material
    (as compared to proteins)
  • bacterial genetics was a new field of study did
    bacteria even contain genes?

12
Discovery of DNA as genetic material
  • Studies using viral DNA (Alfred Hershey Martha
    Chase- 1952)
  • Hershey-Chase experiment
  • involved a bacteriophage T2
  • virus that infects bacteria
  • consists of DNA contained within a protein coat

13
Discovery of DNA as genetic material
  • T2 bacteriophage lifecycle
  • bacteriophage attacks bacterial cell
  • only one portion enters the cell (DNA or
    protein?)
  • 20 minutes later bacterial cell bursts releasing
    dozens of new viruses
  • Bacterial cell gets converted into a
    bacteriophage producing factory.

14
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15
Discovery of DNA as genetic material
  • Hershey-Chase experiment
  • traced the protein and DNA components of the T2
    bacteriophage by radioactively labeling the
    following portions
  • In amino acids cysteine methionine, sulfur is
    a component ? S35
  • In DNA, phosphate is present ? P32

16
Discovery of DNA as genetic material
  • Hershey Chase allowed either the P32 or S35
    labeled viruses to attach to the bacteria
  • After a few minutes, the mixtures were agitated
    which stripped away the parts of the virus that
    were not attached separated this from the
    bacteria
  • Result Most of the S35 labeled portion of the
    bacteriophages seperated from the bacteria

17
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18
Discovery of DNA as genetic material
  • Hershey-Chase Experiment Conclusions
  • DNA was transferred into the bacteria ?
  • DNA is the carrier of genetic information

19
Structure of DNA
  • Things to consider for scientists
  • How is DNA replicated?
  • How does DNA cause the synthesis of specific
    proteins?

20
Structure of DNA- its characterization
  • 1950s Rosalind Franklin attempted to visualize
    the structure of DNA via X-ray crystallographs
  • Revealed a helical structure

21
Structure of DNA- its characterization
  • Erwin Chargaff (1950) found that DNA from many
    different species have equal amounts of the
    nitrogen bases adenine and thymine (AT), and
    guanine and cytosine (GC).
  • Chargaffs rule the abundance of purines (AG)
    equals the amount of pyrimidines (TC)

22
Structure of DNA- its characterization
  • James Watson Francis Crick (1953) built a
    model out of tin that established the general
    structure of DNA
  • Looked at X-ray crystallography results
    determined helical nature, as well as, distances
    within the helix
  • Results of previous density measurements
    suggested that DNA contained 2 strands
  • Previous modeling studies also suggested that the
    2 strands run in opposite directions
    (antiparallel)

23
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24
DNA structure- 5 points
  • DNA is a double-stranded helix
  • DNA has a uniform diameter
  • DNA has a right-handed twist
  • DNA has antiparallel sides
  • The sugar-phosphate backbone coil around the
    outside of the helix, and the nitrogenous bases
    point toward the center.

25
DNA structure- nitrogenous bases
  • Two strands of DNA are held together by hydrogen
    bonds between the nitrogenous bases
  • Adenine (A) pairs with thiamine (T) forming two
    hydrogen bonds
  • Guanine (G) pairs with cytosine (C) by forming
    three hydrogen bonds
  • Complementary Base Pairing

26
DNA structure- antiparallel sides
  • Direction of the polynucleotide can be defined by
    the direction of the phosphodiester linkages
    between adjacent nucleotides
  • diester two bonds formed by OH groups on the
    deoxyribose and phosphate groups
  • notice the position of the 3 and 5 carbons

27
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28
DNA structure- antiparallel sides
  • the two ends of the DNA chain differ
  • one end is a free 5 phosphate group 5end
  • other end is a free 3 hydroxyl group (-OH) 3
    end

29
DNA structure essential to function
  • DNA structure proposed by Watson Crick accounts
    for important functions

30
DNA structure essential to function
  • the nitrogenous base sequence accounts for an
    organisms genetic information ? genetic
    variation
  • genetic material is susceptible to mutation by
    altering of nitrogenous base sequence
  • genetic material can be precisely replicated due
    to complementary base pairing
  • nitrogenous base sequence can be expressed as
    phenotypes in organisms (DNA ? RNA ? protein)

31
How does DNA replicate?
  • Watson-Crick model of DNA suggested that the
    basis for copying genetic information is
    complimentary
  • If
  • 5 ATTGCAT- 3
  • Then the partner sequence is
  • 3 TAACGTA- 5

32
How does DNA replicate?
  • occurs in two steps
  • 1- DNA is unwound to separate the two template
    strands
  • 2- new nucleotides are linked by covalent
    bonding to each new strand in a complementary
    sequence to the old strands

33
Semiconservative replication
  • the two copies of DNA that result from
    replication each contain
  • One newly formed strand of DNA and one old
    strand of DNA

34
How does DNA replicate?
  • new nucleotides are always added to the 3 end of
    DNA
  • there is a free OH group that can react with the
    triphosphate end (or 5 end) of the new
    nucleotide
  • two template strands are replicated, then, in
    opposite directions
  • involves many enzymes, but DNA polymerase is the
    most noteworthy (adds new nucleotides to growing
    strands)

35
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36
FIG. 11.10
37
The replication complex
  • this is a large protein complex that causes the
    interaction of the template DNA and the enzymes
    involved in replication
  • replication complexes attach to various origins
    of replication in the DNA at the same time
  • DNA replicates in both directions from the point
    of origin, creating two replication forks

38
The replication complex
  • Because the DNA strands are antiparalell, the new
    DNA is made in opposite directions
  • DNA going toward the replication fork is made by
    continuously adding new nucleotides (leading
    strand)
  • DNA being made away from the replication fork
    (lagging strand) is synthesized in short segments
    (Okazaki fragments) and are later connected by
    the enzyme ligase .

39
FIG. 11.11
40
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41
The replication complex
  • the replication complexes stay stationary, while
    the template DNA strands move through them

42
Role of DNA polymerase
  • job is to attach new nucleotides to the 3
    growing end of new DNA strands
  • require a RNA primer to attach to DNA (made by
    primase)
  • 3 main protein subunits
  • - large a subunit attaches nucleotides in the
    5? 3 direction
  • - smaller e subunit that proofreads newly formed
    DNA
  • - b2 subunit that clamps the enzyme to the
    template strand

43
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44
Role of DNA polymerase
  • enzyme threads the DNA through the replication
    complex at a rapid rate
  • approx. 1000 nucleotides/second

45
Steps to DNA replication
  • 1. DNA helicase (enzyme) unwinds the DNA. The
    junction between the unwound part and the open
    part is called a replication fork.
  • 2. DNA polymerase adds the complementary
    nucleotides and binds the sugars and phosphates.
    DNA polymerase travels from the 3' to the 5' end.

46
Steps to DNA replication
  • 3. DNA polymerase adds complementary nucleotides
    on the other side of the ladder. Traveling in the
    opposite direction.
  • 4. One side is the leading strand - it follows
    the helicase as it unwinds.
  • 5. The other side is the lagging strand - its
    moving away from the helicase

47
Steps to DNA replication
  • Problem it reaches the replication fork, but the
    helicase is moving in the opposite direction. It
    stops, and another polymerase binds farther down
    the chain.
  • This process creates several fragments, called
    Okazaki Fragments, that are bound together by DNA
    ligase.

48
Steps to DNA replication
  • 6. During replication, there are many points
    along the DNA that are synthesized at the same
    time (multiple replication forks). It would take
    forever to go from one end to the other, it is
    more efficient to open up several points at one
    time.

49
An animation
  • http//www.stolaf.edu/people/giannini/flashanimat/
    molgenetics/dna-rna2.swf

50
DNA proofreading and repair
  • not doing this can have a big price
  • the incorrect transfer of genetic information to
    new cells

51
3 DNA repair systems
  • 1- proofreading corrects errors in replication
    as DNA polymerase makes new strands
  • 2- mismatch repair DNA is scanned immediately
    after it is made and base-pairing mismatches are
    corrected
  • 3- excision repair abnormal N- bases are removed
    due to chemical damage, and replace with
    functional N- bases.
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