The Molecular Basis of Inheritance

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The Molecular Basis of Inheritance

• Chapter 16

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HISTORY OF DNA STRUCTURE

• How did we learn that DNA is the key to coding
  for all characteristics of living things?
• Once Morgans group showed that genes are located
  on chromosomes, the two chemical components of
  chromosomes DNA PROTEIN, became the
  candidates for the genetic material.

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TIMELINE

• 1928
• British scientist -- Frederick Griffith studies
  bacteria looking for cause of pneumonia
• found two specific strains or cultures of
  bacteria that looked different when growing on
  petri dishes
• one grew in smooth-edged groups
• other one produced colonies that were rough and
  ragged around the edges
• IMPORTANCE
• Visual differences made it easy to recognize and
  distinguish between the strains of bacteria
• Also, Griffith found that
• smooth-edged colonies of bacteria caused disease
• rough-edged colonies were harmless

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Griffiths Experimenthttp//nortonbooks.com/colle
ge/biology/animations/ch12a01.htm
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RESULTS OF GRIFFITHS 1928 EXPERIMENT

• Discovery of process of transformation
• Somehow the heat-killed bacteria had passed their
  disease-causing ability to the harmless strain
• The harmless strain had been transformed into a
  disease-causing strain
• Hypothesized that some factor was responsible
  for this change

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TIMELINE CONTINUED

• 1944
• American, Oswald Avery, continued bacteria
  research of Griffith
• Knew were 4 types of organic compounds that make
  up all life
• used enzymes to destroy lipids, carbohydrates,
  proteins, and RNA in an extract from the disease
  causing bacteria.
• IMPORTANCE
• Transformation still occurred, so obviously the
  molecules they had destroyed were not responsible
  for transformation.
• Only organic molecule left that had not been
  destroyed was DNA
• When repeated experiment with DNA-destroying
  enzymes, no transformation occurred.DNA was the
  key to heredity

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TIMELINE CONTINUED

• 1952
• Americans Alfred Hershey and Martha Chase
• worked with viruses called bacteriophages
• viruses are simple DNA or RNA core and a
  protein coat around them
• when they infect, bacteriophages inject DNA or
  RNA into cell and protein coat is left outside
• used radioactive markers to trace
• phosphorus-32 (32P) for DNA
• sulfur-35 (35S) for protein coat

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Figure 16.2a The Hershey-Chase Experiment Phages
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Hershey-Chase Experimenthttp//nortonbooks.com/co
llege/biology/animations/ch12a02.htm
Bacteriophage with phosphorus-32 in DNA
Phage infectsbacterium
Radioactivity inside bacterium
Bacteriophage with sulfur-35 in protein coat
Phage infectsbacterium
No radioactivity inside bacterium
Go to Section
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RESULTS OF HERSHEY-CHASE

• When viruses were separated from the bacteria and
  tested for radioactivity, all of the
  radioactivity from the bacteria was found to be
  32P
• Conclusion
• genetic material of the bacteriophage that was
  transferred was DNA NOT PROTEIN!

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RACE FOR STRUCTURE OF DNA

• 1940
• Erwin Chargaff discovers that percentages of A
  and T are equal in any sample of DNA same is
  true for C and G
• 1944
• Linus Pauling discovers that proteins can have a
  helical shape
• 1952
• Rosalind Franklin takes pictures of DNA molecule
  using technique called X-ray diffraction, shows
  that DNA has helical shape

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Figure 16.4 Rosalind Franklin and her X-ray
diffraction photo of DNA
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RACE FOR STRUCTURE OF DNA

• 1951-1952
• Maurice Wilkins works with X- ray diffraction and
  sees same pattern as Franklin, shares info with
  James Watson
• April, 1953
• James Watson and Francis Crick build first model
  of DNA (are awarded Nobel Prize in 1960s)

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Figure 5.x3 James Watson and Francis Crick
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Meselson-Stahl Experiment

• Late 1950s experiments by Meselson Stahl
  support the semi-conservative replication of DNA
• Data reject conservative and dispersal hypotheses!

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Practice Questions

• Which of the following is LEAST related to the
  others?
• Transformation
• Phage
• DNA
• Griffeth
• Avery
• Answer phage

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Practice Questions

• What was the contribution of the following
  scientists regarding the discovery of our present
  knowledge of the nature of genes and/or the shape
  of the DNA?
• Griffith
• Hershey Chase
• Avery, MacLeod, and McCarty
• Chargaff
• Meselson Stahl

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BASIC DNA STRUCTURE

• Exists as a double helix
• Backbone of DNA structure made up of alternating
  sugar (deoxyribose) and phosphate groups
• Bases are attached to the sugars
• Bases in DNA are adenine, thymine, cytosine, and
  guanine
• A pairs with T, C pairs with G and vice-versa
• A and G are purines larger, double rings
• T and C are pyrimidines smaller, single rings

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Figure 16.6 Base pairing in DNA
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Why does A always pair with T (or U), and G with
C?
Distance between uprights is 2 nm (nanometers)
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Chromosome Structure of Eukaryotes
Eukaryotic chromosomes contain DNA wrapped around
proteins called histones. The strands of
nucleosomes are tightly coiled and supercoiled to
form chromosomes.
Go to Section
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3 Important Chromosome Functions

• Carry information from one generation to the
  next.
• Put that information to work by determining the
  heritable characteristics of organisms.
• Have to be easily copied each time a cell
  divides.

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Practice Question

• All of the following elements are present in DNA
  except
• Oxygen
• Nitrogen
• Carbon
• Sulfur
• Phosphorus
• Answer sulfur

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Practice Questions

• Cytosine makes up 38 of the nucleotides in a
  sample of DNA from an organism. What percent of
  the nucleotides in this sample will be thymine?
• 12
• 24
• 31
• 38
• Cannot be determined with information provided
• Answer 12

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Practice Questions

• In an analysis of the nucleotide composition of
  DNA, which of the following is TRUE?
• A C
• A G and C T
• A C G T
• A T G C
• Both B and C are true
• Answer A C G T

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Practice Question

• All of the following were determined directly
  from X-ray diffraction photographs of
  crystallized DNA except
• The diameter of the double helix
• The helical shape of DNA
• The sequence of nucleotides
• The linear distance required for one full turn of
  the double helix
• The width of the double helix
• Answer the sequence of nucleotides

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DNA REPLICATION

• Open up the helix to promote easier copying
  HELICASE is the enzyme that facilitates this
• There are MANY helicases that work together to
  unwind the double helix efficiently.
• WATCH THIS ANIMATION SEVERAL TIMES
• http//www.wiley.com/college/pratt/0471393878/stud
  ent/animations/dna_replication/index.html

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Figure 16.7 A model for DNA replication the
basic concept (Layer 1)
REMEMBER this all occurs during the S phase of
interphase in mitosis or- during MEIOSIS I ONLY!
http//bcs.whfreeman.com/thelifewire/content/chp11
/1102002.html http//bcs.whfreeman.com/thelifewir
e/content/chp11/1102002.html http//bcs.whfreeman
.com/thelifewire/content/chp11/1102003.html
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Figure 16.7 A model for DNA replication the
basic concept (Layer 2)
Hydrogen bonds are broken here!!!
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Figure 16.7 A model for DNA replication the
basic concept (Layer 3)
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Figure 16.7 A model for DNA replication the
basic concept (Layer 4)
SEMI-CONSERVATIVE PROCESSwhat does this mean?
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Semi-Conservative Replicationhttp//www.sumanasin
c.com/webcontent/animations/content/meselson.html
The 2 strands of the parental molecule separate,
and each functions as a template for synthesis of
a new complimentary strand. SEE TEXT PAGE 294.
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Replication

• The replication of a DNA molecule begins at an
  origin of replication.
• Bacterial chromosomes are circular and have a
  SINGLE origin of replication.
• Eukaryotic chromosomes are linear and may have
  hundreds of origins of replication.
• Proteins that initiate DNA replication recognize
  these sequences and attach to the DNA, separating
  the two strands and opening up a replication
  bubble.
• DNA replication occurs in a Replication Bubble
  and each end is called a Replication fork
• Replication is catalyzed by the enzyme DNA
  polymerase makes sure that correct bases pair.
• Llike a spell check for the process of
  replication.
• DNA Ligase is the enzyme that forms the new
  covalent bonds between the DNA nucleotides.

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Replication begins at specific sites where the
two parental strands separate to form replication
bubbles. The bubbles expand laterally, as DNA
replication proceeds in both directions. Eventual
ly, the replication bubbles fuse, and synthesis
of the daughter strands is complete.
REPLICATION FORK a y-shaped region where new
strands of DNA are elongating.
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Figure 16.11 Incorporation of a nucleotide into
a DNA strand
Where does the energy for DNA replication come
from?
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Figure 16.12 The two strands of DNA are
antiparallel
The two DNA strands are ANTIPARALLEL that is,
their sugar-phosphate backbones run in opposite
directions. The 5 ? 3 direction of one strand
runs counter to the 5 ? 3 direction of the
other strand. The numbers assigned to the carbon
atoms of the deoxyribose are shown for two of
them. In the figure, the five carbons of one
deoxyribose sugar of each DNA strand are numbered
from 1 to 5. Notice in the figure that a
nucleotides phosphate group is attached to the
5 carbon of deoxyribose. Notice also that the
phosphate group of one nucleotide is joined to
the 3 carbon of the adjacent nucleotide.
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Antiparallel Arrangement of DNA

• Replicating the LEADING STRAND
• Antiparallel sugar-phosphate backbones run in
  opposite directions
• See page 296 in text
• DNA polymerases add nucleotides ONLY to the free
  3 end of a growing DNA strand, never to the 5
  end
• SO, a new DNA strand can only elongate in the 5?
  3 direction!
• Polymerase nestles in the replication fork on the
  template strand that is read 5 ? 3 (leading
  strand)
• This strand is replicated continuously in the 5
  to 3 direction.

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Antiparallel Arrangement of DNA

• Replicating the LAGGING STRAND
• To elongate the other new DNA strand, polymerase
  must work along the other template strand in the
  direction AWAY FROM the replication fork.
• As a replication bubble opens, a polymerase
  molecule can work its way away from a replication
  fork and synthesize a short segment of DNA.
• As the bubble grows, another short segment of the
  lagging strand can be made in a similar way.
• This is known as DISCONTINUOUS replication
  because the DNA is synthesized in short fragments
  called OKAZAKI fragments.

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Practice Question

• The strands that make up DNA are antiparallel.
  What does this mean?
• Answer the 5 to 3 direction of one strand runs
  counter to the 5 to 3 direction of the other
  strand

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Figure 16.13 Synthesis of leading and lagging
strands during DNA replication
1. DNA polymerase elongates DNA strands only in
the 5 ? 3 direction.
2. One new strand (leading strand) can therefore
elongate continuously 5 ? 3 as the replication
for progresses.
3. The other new strand (lagging strand), must
grow in an overall 3 ? 5 direction by addition
of short segments (Okazaki fragments) that grow
5 ? 3 (numbered here in the order they were
made).
4. Ligase connects the Okazaki fragments.
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Priming DNA Synthesis

• DNA polymerases cannot initiate synthesis of a
  polynucleotide it can only add nucleotides to
  already existing chain!
• The start of a NEW chain is not DNA, but a short
  stretch of RNA called a primer.
• The enzyme primase joins this RNA to the DNA
  template to make the primer.
• Primase can start an RNA chain from scratch!

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Figure 16.14 Priming DNA synthesis with RNA
DNA polymerase cannot initiate a polynucleotide
strand it can only add to the 3 end of an
already-started strand. The primer is a short
segment of RNA synthesized by the enzyme primase.
Each primer is eventually replaced by DNA.
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Figure 16.15 The main proteins of DNA
replication and their functions
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Figure 16.16 A summary of DNA replication
http//highered.mcgraw-hill.com/olc/dl/120076/bio2
3.swf
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Practice Question

• Which enzymes catalyze the elongation of a DNA
  strand in the 5 to 3 direction?
• Primase
• DNA ligase
• DNA polymerases
• Topoisomerase
• Helicase
• Answer DNA polymerases

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Practice Question

• What is the function of the following enzymes in
  DNA replication?
• Helicase
• SSBs
• Nuclease
• Ligase
• DNA polymerase
• primase

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Practice Question

• In DNA, the designations 3 and 5 refer to what?
• Answer carbon atoms of deoxyribose to which
  phosphate groups may bond

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Practice Question

• Which of the following is LEAST related to the
  others on the list?
• Okazaki fragment
• Primer
• Telomere
• Leading strand
• Lagging strand
• Answer telomere

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Practice Question

• If I am writing an essay about DNA replication
  what pertinent vocabulary terms should I use AND
  define?

Antiparallel arrangement Helicase Hydrogen bonds SSBs
Base pairing Semi-conservative Replication bubble Replication fork
DNA polymerase Primase RNase H Ligase
RNA primer 5 to 3 Leading strand (continuous) Lagging strand (discontinuous)
Okazaki fragments complementary S phase of mitosis Meiosis I
nuclease Telomeres telomerase
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PROOFREADING DNA FOR ERRORS

• Enzymes proofread DNA during its replication and
  repair damage in existing DNA
• See page 299
• errors in completed DNA molecule amount to only
  one in a billion nucleotides
• Initial pairing errors error rate of 1 per
  10000 base pairs
• exposure damage reactive chemicals, radiation,
  X-rays, ultraviolet light (unpredictable, but
  common)
• Ex. Skin cells and UV damage
• Mismatch repair cells use special enzymes to
  fix incorrectly paired nucleotide
• Nucleotide excision repair uses nuclease (DNA
  cutting enzyme that excises damaged area)

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Figure 16.17 Nucleotide excision repair of DNA
damage
A team of enzymes detects and repairs damaged
DNA. Repair enzymes can excise damaged DNA
regions from the DNA and replace them with a
normal segment. Good Animation http//nortonbook
s.com/college/biology/animations/ch12a05.htm
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The Dilemma of DNA Ends

• For linear DNA, like that found in eukaryotes,
  DNA polymerase can only add nucleotides to the 3
  end of a preexisting polynucleotide this is a
  potential problem!
• The normal replication machinery has no way to
  complete the 5 ends of daughter DNA strands as
  a result repeated rounds of replication produce
  shorter and shorter DNA molecules.
• Result deletion of essential genes!
• NOTE prokaryotes avoid this problem by having
  circular DNA!

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Figure 16.18 The end-replication problem
When a linear DNA molecule replicates, a gap is
left at the 5 end of each new strand (light
blue) because DNA polymerase can only add
nucleotides to a 3 end. As a result, with
each round of replication, the DNA molecules get
slightly shorter. Good Animation http//spine.ru
tgers.edu/cellbio/assets/flash/tel.htm
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Telomeres and Telomerase

• Eukaryotic chromosomal DNA molecules have special
  nucleotide sequences called telomeres at their
  ends
• Telomeres are regions of DNA that do not contain
  genes rather, are multiple repetitions of one
  short nucleotide sequence.
• Typically, the repeated unit TTAGGG is seen in
  human telomeres.
• Telomeric DNA protects an organisms genes from
  being eroded through successive rounds of DNA
  replication!
• These (along with special protein called
  telomerase associated with it) prevent the ends
  from activating the cells systems for monitoring
  DNA damage.
• Ex. The end of DNA strand seen as double strand
  break might trigger signal transduction
  pathways leading to cell cycle arrest or cell
  death)!

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Figure 16.19b Telomeres and Telomerase
SOME eukaryotes deal with the end-replication
issues by having expendable, noncoding sequences
called telomeres at the ends of their DNA and the
enzyme telomerase in some of their cells. In the
long term, over the course of generations,
eukaryotic organisms need a way of restoring
their shortened telomeres. This is provided by
telomerase (a special enzyme that catalyzes the
lengthening of telomeres).
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Telomeres Limiting Life?

• Telomerase is NOT present in most cells of
  multicellular organisms like ourselves, and the
  DNA of dividing somatic cells does tend to be
  shorter in older individuals and in cultured
  cells that have divided many times.
• Thus, it is possible that telomeres are a
  limiting factor in the life span of certain
  tissues and even organisms as a whole.
• Telomerase is however present in germ-line cells
  that give rise to gametes and here the enzyme
  produces long telomeres in these cells and hence
  in the newborn.
• Intriguingly, telomerase is also found in somatic
  cells that are cancerous these usually have
  unusually short telomeres, which one would expect
  for cells that have undergone many rounds of
  division.
• Progressive shortening would eventually lead to
  self-destruction of cancer unless telomerase
  became available to stabilize telomere length.
• This is exactly what seems to happen in cancer
  cells. IF this is an important factor it may
  well provide a useful target for cancer diagnosis
  and chemotherapy!

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The Essay

• Scientists seeking to determine which molecule is
  responsible for the transmission of
  characteristics from one generation to the next
  knew that the molecule must
• (1) copy itself precisely,
• (2) be stable but able to be changed, and
• (3) be complex enough to determine the organism's
  phenotype.
• Explain how DNA meets each of the three criteria
  stated above.
• Select one of the criteria stated above and
  describe experimental evidence used to determine
  that DNA is the hereditary material.