Lecture 9
DNA Structure & Replication
What is a Gene?
Mendel’s work left a key question unanswered:
What is a gene?
The work of Sutton and Morgan established that
genes reside on chromosomes
But chromosomes contain proteins and DNA
So which one is the hereditary material
Several experiments ultimately revealed the
nature of the genetic material
The Griffith Experiment
In 1928, Frederick Griffith discovered transformation
while working on Streptococcus pneumoniae
The bacterium exists in two strains
S
Forms smooth colonies in a culture dish
Cells produce a polysaccharide coat and can cause disease
R
Forms rough colonies in a culture dish
Cells do not produce a polysaccharide coat and are therefore
harmless
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How Griffith Discovered Transformation
Thus, the dead S bacteria
somehow “transformed” the live R
bacteria into live S bacteria
The Avery and Hershey-Chase Experiments
Two key experiments that demonstrated conclusively
that DNA, and not protein, is the hereditary material
Oswald Avery and his coworkers Colin MacLeod and
Maclyn McCarty published their results in 1944
Alfred Hershey and Martha Chase published their
results in 1952
The Avery Experiments
Avery and his colleagues prepared the same mixture of dead
S and live R bacteria as Griffith did
They then subjected it to various experiments
All of the experiments revealed that the properties of the
transforming principle resembled those of DNA
1.
2.
3.
4.
Same chemistry and physical properties as DNA
Not affected by lipid and protein extraction
Not destroyed by protein- or RNA-digesting enzymes
Destroyed by DNA-digesting enzymes
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The Hershey-Chase Experiment
Viruses that infect
bacteria have a simple
structure
DNA core surrounded
by a protein coat
Hershey and Chase used
two different radioactive
isotopes to label the protein
and DNA
Incubation of the labeled
viruses with host bacteria
revealed that only the DNA
entered the cell
Therefore, DNA is the
genetic material
Thus, viral DNA
directs the
production of new
viruses
Discovering the Structure of DNA
Chemically, DNA is made up of nucleotides
Each nucleotide has a central sugar, a phosphate group and an
organic base
The bases are of two
main types:
Nitrogenous
base
Purines
Large bases:
Adenine (A)
Guanine (G)
5-C
sugar
Pyrimidines
Small bases:
Cytosine (C)
Thymine (T)
More Key Observations About DNA
Erwin Chargaff made key DNA observations that became known as
Chargaff’s rule
Purines = Pyrimidines
A=T
C=G
Rosalind Franklin’s X-ray
diffraction experiments revealed
that DNA had the shape of a
coiled spring or helix
Rosalind
Franklin (19201958)
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The Structure of DNA Revealed
In 1953, James Watson and Francis Crick deduced that DNA was a
double helix
They came to their conclusion using Tinker toy models and the research of
Chargaff and Franklin
James Watson
(1928)
Francis Crick
(1916-2004)
The DNA Double Helix
The two DNA strands are held
together by weak hydrogen bonds
between complementary base pairs
Dimensions
suggested by
X-ray diffraction
A and T
C and G
If the sequence on one strand is
ATACGCAT
TATGCGTA
The other’s sequence must be
The two
possible
base pairs
Each chain is a complementary
mirror image of the other
So either can be used as template
to reconstruct the other
3 possible methods for DNA replication
Daughter DNAs
contain one old
and one new
strand
Original DNA
molecule is
preserved
Old and new
DNA are
dispersed in
daughter
molecules
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Evidence for Semi-Conservative Replication
These three mechanisms were tested
in 1958 by Matthew Meselson and
Franklin Stahl
Thus, DNA
replication is semiconservative
Overview of How DNA Copies Itself
DNA helices begin unwinding
from the nucleosomes
Helicase untwists the double
helix and exposes
complementary strands
The enzyme primase puts
down a short piece of RNA
termed the primer
DNA polymerase reads along
each naked single strand
adding the complementary
nucleotide
The site of replication is the
replication bubble
Each nucleotide strand serves
as a template for building a
new complementary strand
DNA Replication Overview
PLAY
How nucleotides are added in DNA replication
Template strand
HO 3’
Sugarphosphate
backbone
New strand
O
P
T
A
T
P
O
A
P
O
O
P
T
A
DNA polymerase
C
G
P
O
O
O
P
C
P
O
O
T
A
P
O
G
P
O
O
P
A
3’
OH
A
O
T
O
O
P
P P
P
Pyrophosphate
P
A
O
T
O
A
P P P
P
5’
P
G
O
P
P
5’
C
O
O
P
New strand
HO 3’
P
G
O
P
Template strand
5’
C
P
OH
3’
OH
O
5’
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Summary of How DNA Copies Itself
The process of DNA
replication can be
summarized as such
The enzyme helicase first
unwinds the double helix
The enzyme primase puts
down a short piece of RNA
termed the primer
DNA polymerase reads
along each naked single
strand adding the
complementary nucleotide
DNA polymerase can only build a strand of DNA in one direction
The leading strand is made continuously from one primer
The lagging strand is assembled in segments created from many primers
DNA Replication
Since DNA polymerase
only works in one direction:
A continuous leading
strand is synthesized
A discontinuous lagging
strand is synthesized
DNA ligase splices
together the short
segments of the
discontinuous strand
Two new telomeres are
also synthesized
This process is called
semiconservative
replication
DNA Replication (cont.)
RNA primers are removed and replaced with DNA
Ligase joins the ends of newly-synthesized DNA
Mechanisms exist for DNA proofreading and repair
PLAY
DNA Replication
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Transcription
The path of genetic information is often called the central
dogma
DNA
RNA
Protein
A cell uses three kinds of RNA to make proteins
Messenger RNA (mRNA)
Transfer RNA (tRNA)
Ribosomal RNA (rRNA)
Transcription
Gene expression is the use of information in DNA to direct
the production of proteins
It occurs in two stages
Transcription
The transcriber is
RNA polymerase
It binds to one DNA strand at a site
called the promoter
It then moves along the DNA
pairing complementary nucleotides
It disengages at a stop signal
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Working with DNA
Key techniques used by today’s genetic engineers include
PCR amplification
Used to increase the amounts of DNA
cDNA formation
Used to build genes from their mRNA
DNA fingerprinting
Used to identify particular individuals
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