Kwame Nkrumah University of

Science & Technology, Kumasi, Ghana

Module 2

DNA Replication
Prof. Matthew Glover Addo

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Module 2 Module Dashboard FILTER

DNA Replication
Lessons Assignments
4 Start Learning
3
Dashboard
hig

Pick a Lesson
Introduction
Lesson 1

Method of
t1

Lessons
h1

title
Replication
Assignments Lesson 2
t2

Mechanism of
Play
h2

Replication
Watch an overview
Lesson 3
video on DNA t3

Rate of Replication
h3

2
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Module 2 Module Objectives

DNA Replication

Dashboard 1 Consider the modes of replication i.e. dispersive,

conservative and semiconservative
Introduction
hig

Lessons
2 Consider the experiments of Meselson and Stahl
title

Assignments
Their experiment gave a convincing evidence to support
the semiconservative nature of replication

3
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Module 2 Module Objectives

DNA Replication

Dashboard
3 To appreciate how replication is initiated,
elongated and terminated
Introduction
Emphasis will be laid on some of the enzymes
hig

Lessons involved in these processes

title

Assignments
4 DNA repair mechanism

4
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Module 2 Module Lessons

DNA Replication

Dashboard

Introduction
t1 t2 t3

Lessons
hig

Lesson 1 h1
Lesson 2 title
h2
Lesson 3 h3

Assignments
Method of Mechanism of Rate of
Replication Replication Replication

Start Start Start

5
 Module 2

DNA Replication

t1
h1

Lesson 1
hig

Method of DNA Replication t3

Learn about the three hypotheses that were previously

proposed for the method of replication of DNA
t2

Learn Now
 Lesson 1

Method of DNA Replication t1

FILTER

To explain the
phenomenon of
heredity, biological
information must
hig

be accurately h1

copied (replicated)
and transmitted
from each cell to all
of its progeny 7
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses t1
FILTER

Conservative Dispersive Semiconservative

hig

h1

1 2 3

8
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Conservative
t1
FILTER

Conservative replication
would leave intact the original
DNA molecule and generate a
hig

h1
1

completely new molecule

9
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Dispersive
t1
FILTER

Dispersive replication would

produce two DNA molecules
with sections of both old and
hig
2

h1

new DNA interspersed along

each strand

10
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Dispersive
t1
FILTER

The dispersive hypothesis is

exemplified by a model
proposed by Max Delbrück
hig

h1 2

Learn More

11
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Dispersive
t1
FILTER

▪ He attempted to solve the

problem of unwinding the
two strands of the double
helix by a mechanism that
hig

breaks the DNA backbone h1

2

every 10 nucleotides

▪ Untwist the molecule, and

attaches the old strand to 12
t3
t2

the newly synthesized one

 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Dispersive
t1
FILTER

This would synthesize the

DNA in short pieces
alternating from one
hig
2

h1

strand to the other

13
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER

Semiconservative replication
would produce molecules
with both old and new DNA,
but each molecule would be
hig

3
h1

composed of one old strand

and one new one

14
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER

Each strand acts as

a template or
guide for the
synthesis of a new
hig

DNA molecule by h1
3

the sequential
addition of
complementary
base pairs 15
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER

Thereby generating
a new DNA strand
that is the
complementary
hig

3
h1

sequence to the
parental DNA

16
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER

Each daughter DNA

molecule ends up with
hig

one of the original h1

strands and one newly

3

synthesized strand

17
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment
In 1958
Matthew Meselson
& Franklin Stahl
worked out a clever
hig

procedure to distinguish h1
3

semi conservative DNA

replication from conservative
or dispersive replication,
using a nonradioactive heavy
isotope of nitrogen 18
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

Meselson and Stahl

opted for nitrogen
hig

because it is an h1
3

essential chemical
component of DNA

19
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment
Therefore, every time a cell divides and its DNA replicates, it
incorporates new N atoms into the DNA of either one or both of
its two daughter cells, depending on which model was correct

hig

h1

20
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

Ordinary nitrogen,
the most abundant
hig
isotope, has an 3

atomic weight of 14,

h1

so it is called 14N

21
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

A relatively rare
isotope 15N has
hig
an atomic weight 3

of 15
h1

22
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

Meselson and Stahl

found that if bacteria
are grown in a medium
enriched in 15N, they
incorporate the heavy
hig 3

h1

isotope into their DNA,

which becomes denser
than normal
23
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

This labeled DNA

clearly separates
from ordinary DNA
hig

in gradient of h1
3

Cesium Chloride
(CsCl) spun in an
ultracentrifuge
24
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

CsCl is used because

it is a very dense salt
and therefore makes
dense enough
solution that DNA
hig 3

h1

will float somewhere

in the middle rather
than sinking to the
bottom 25
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

The aim of the

experiment was to
grow 15N labeled
bacteria in 14N-
hig 3

h1

medium and then to

look at the density of
the DNA products
26
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

That is, E. coli cells

with only 15N in
their DNA were
transferred to a 14N
hig 3

h1

medium and were

allowed to divide or
replicate
27
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

The progress of
cell division was
monitored by
measuring the
hig 3

h1

optical density of
the cell suspension

28
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

DNA was extracted

periodically and was
compared to pure 14N
hig 3

h1

DNA and 15N DNA

29
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results
Based on these findings, the scientists were immediately able to
exclude the conservative model of replication as a possibility

hig

h1

1 2 3

30
Conservative Dispersive Semiconservative
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

After all, if DNA

replicated conservatively,
there should have been
two distinct bands after a
1

hig

h1

single round of
replication

31
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

However, this result

was consistent with
both semiconservative
2 3

hig

h1

and dispersive
replication

Dispersive Semiconservative 32
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Experiment

To differentiate
between the two,
Meselson and Stahl
hig
had to let the cells 3

divide again and then

h1

sample the DNA after

a second round of
replication
33
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

DNA from cells after

two replications had
been completed was
hig

found to consist of h1
3

equal amounts of
DNA with two
different densities
34
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results
Intermediate density
of DNA of cells grown DNA from cells
for only one division in grown exclusively
14N medium in 14N medium
hig

h1

3 3

35
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

This was inconsistent

with dispersive
replication
3

After all, if the dispersive

hig

h1

model were the correct model,

the scientists would have
continued to observe only a
single band after every round
of replication 36
t3
t2

Dispersive Semiconservative
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results
Dispersive replication
would have resulted in
double-stranded DNA
with both strands
hig

having mixtures of 15N h1

and 14N DNA, either of

2

which would have

appeared as DNA of an
intermediate density Dispersive
37
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

Semiconservative
replication would
result in double-
stranded DNA with
3

hig

h1

one strand of 15N

DNA, and one of
14N DNA
Semiconservative
38
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

This will result in equal

hig
amounts of DNA with h1
3

two different densities

Semiconservative 39
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results

The scientists continued to

observe the same two bands
after several subsequent
3

hig

h1

rounds of replication

Semiconservative 40
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results
These results were
consistent with the
semiconservative model
of replication and the
hig

reality that, when DNA h1

3

replicated, each new

double helix was built
with one old strand and
one new strand Semiconservative 41
t3
t2
 Lesson 1
Method of DNA
Replication
Proposed Hypotheses Semiconservative
t1
FILTER Results
The result was consistent with the semiconservative
replication hypothesis

hig

3
h1

1 2

42
Conservative Dispersive Semiconservative
t3
t2
 Lesson 1
Method of DNA
Replication
Replication of Eukaryotic Chromosomes
t1
FILTER

Before a cell can

divide, it must
hig

duplicate or h1

replicate all of
its DNA

43
t3
t2
 Lesson 1
Method of DNA
Replication
Replication of Eukaryotic Chromosomes
t1
FILTER

In eukaryotes, G2 Cell Division

this occurs S M
hig

during S phase of h1

Interphase
g2
g1
mi
mcsa
p
it

the cell cycle

G1

44
t3
t2
 Lesson 1
Method of DNA
Replication
Replication of Eukaryotic Chromosomes
t1
FILTER

Eukaryotic DNA
replication is very
slow compared
hig
to E. coli DNA
replication: only
h1

about 75
nucleotides/
second
45
t3
t2
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FILTER

Module 2 Module Lessons

DNA Replication

Dashboard

Introduction
t1 t2 t3

Lessons
hig

Lesson 1 h1
Lesson 2 title

h2
Lesson 3 h3

Assignments
Method of Mechanism of Rate of
Replication Replication Replication

Start Start Start

46
 Module 2

DNA Replication

Lesson 2
t2
h2

hig

Mechanism of DNA
Replication t3

Learn about the mechanism DNA uses to

replicate itself to form new DNAs
t1

Learn Now 47
 Lesson 2

Mechanism of DNA Replication

t2
FILTER

Replication is a
huge task, whether
in bacteria or in
hig

eukaryotes and h2

requires many
proteins or
enzymes to act
together 48
t3
t1
 Lesson 2

Mechanism of DNA Replication

Physical & Biochemical Challenges Cells Overcome
t2
FILTER

1 Locating site(s) to
begin replication
The proper enzymes
must be collected
or deposited there
hig

h2

49
t3
t1
 Lesson 2

Mechanism of DNA Replication

Physical & Biochemical Challenges Cells Overcome
t2
FILTER

1 Locating site(s) to
begin replication

2 Unwinding the
hig
double helix
to expose the two
h2

strands. This imposes

twisting strain on the
portions of the helix
farther away from the 50
t3
t1

unwinding site
 Lesson 2

Mechanism of DNA Replication

Physical & Biochemical Challenges Cells Overcome
t2
FILTER

1 Locating site(s) to
begin replication

2 Unwinding the double helix

hig
This is much like how
untangling a twisted
h2

phone cord does,

and those forces
must be relieved to
prevent breakage of 51
t3
t1

the DNA strands

 Lesson 2

Mechanism of DNA Replication

Origins t2
FILTER

DNA replication begins

at specific points known
as the Origins
hig

These sites are

h2

recognized by certain
proteins in the cell

52
t3
t1
 Lesson 2

Mechanism of DNA Replication

Origins t2
FILTER

The Origins are rich in Adenine

and Thymine bases, as breaking
2 hydrogen bonds between the
hig

Adenine and Thymine is easier h2

than breaking the triple

hydrogen bonds between
Cytosine and Guanine
53
t3
t1
 Lesson 2

Mechanism of DNA Replication

Replication Fork t2
FILTER

6 proteins arranged in
a ring shape known as
Helicase, unwind the
double stranded DNA
helix into single
hig

h2

strands by breaking
the hydrogen bonds
between them
54
t3
t1
 Lesson 2

Mechanism of DNA Replication

Replication Fork t2
FILTER

This results in the

formation of a
replication fork
hig

The replication fork is a

h2

structure that is formed

during the DNA
replication process
55
t3
t1
 Lesson 2

Mechanism of DNA Replication

Replication Fork t2
FILTER

The fork is made with

the action of helicase,
that breaks the
hydrogen bonds, that
hig

h2

hold the two DNA

strands together

56
t3
t1
 Lesson 2

Mechanism of DNA Replication

Replication Fork t2
FILTER

This results in a
structure that has
hig

two branching h2

'prongs' of a single
strand DNA each

57
t3
t1
 Lesson 2

Mechanism of DNA Replication

Tetramers t2
FILTER

Tetramers, known as
the single stranded
hig

binding proteins, h2

cover the single-

stranded DNA

58
t3
t1
 Lesson 2

Mechanism of DNA Replication

Tetramers t2
FILTER

This prevents the

DNA strands from
hig

re-annealing and h2

forming the double

stranded molecule

59
t3
t1
 Lesson 2

Mechanism of DNA Replication

The two single DNA strands t2
FILTER

The two single

DNA strands act
as templates
hig
individually, that h2

are used for

producing two
complementary
DNA strands
60
t3
t1
 Lesson 2

Mechanism of DNA Replication

The two single DNA strands t2
FILTER

The double helix

consists of two
anti-parallel DNA
strands with
hig

h2

complementary
5' to 3' strands
61
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

An RNA polymerase
known as primase,
synthesizes short
hig

RNA primers (about h2

60nt long) that

initiate the DNA
replication process
62
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

An enzyme called
DNA polymerase
binds to one
hig
strand of the
DNA and begins
h2

moving along it
in the 3' to 5'
direction
63
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

DNA polymerase
cannot begin
synthesizing the DNA
hig
strand initially h2

It needs a nucleic acid

chain in the beginning
to begin copying the
strand
64
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

The original DNA

strand is used as a
template to
synthesize the DNA
strand in the 5' → 3'
hig

h2

direction with the

help of an extension
formed by RNA
primer 65
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

DNA polymerase
hig

can synthesize the h2

strand in 5' → 3'

direction only

66
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

It hooks the 5' phosphate

group of an incoming
nucleotide onto the 3'
hig

hydroxyl group at the h2

end of the growing

nucleic acid chain

67
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

In eukaryotes,
this molecule is
hig

called DNA h2

polymerase
delta (δ)

68
t3
t1
 Lesson 2

Mechanism of DNA Replication

Initiating the Replication Processt2
FILTER

The DNA
polymerase
uses the strand
as a template
for assembling a
hig

h2

leading strand
of nucleotides
and reforming a
double helix 69
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Leading Strand t2
FILTER

The DNA strand that

is read in the 3' → 5'
and synthesized in
hig

the 5' → 3' direction h2

continuously, is
known as the
leading strand
70
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Leading Strand t2
FILTER

DNA polymerase
III synthesiSes the
hig

DNA using the 3'- h2

OH group, donated
by the single RNA
primer
71
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Leading Strand t2
FILTER

The DNA replication

continues in the
hig

direction of the h2

replication fork, in a
continuous manner

72
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Leading Strand t2
FILTER

RNAse H and DNA

polymerase I
(exonuclease)
recognizes the RNA
hig

polymers that are h2

bound to the DNA

template and
removes the primers
by RNA hydrolysis 73
t3
t1
 t2
FILTER

Module 2 Module Assignments

DNA Replication
Assignment 1

Dashboard

Introduction Read on
Lessons
DNA POL I, POL II
and POL III
t
h2 i
t
l
e

Assignments
hig

74
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Lagging Strand t2
FILTER

The lagging strand is

the DNA strand of
hig

the replication fork, h2

that is opposite to
the leading strand

75
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Lagging Strand t2
FILTER

Because DNA synthesis

can only occur in 5' to 3‘
direction, a molecule of
a second type of DNA
hig

polymerase (epsilon, ε, h2

in eukaryotes) binds to
the other template
strand as the double
helix opens 76
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Lagging Strand t2
FILTER

This molecule
synthesize
discontinuous
hig

segments of h2

polynucleotides
(called Okazaki
fragments)
77
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Lagging Strand t2
FILTER

Primase builds
RNA primers in
short bursts over
the lagging
strand, which is
hig

h2

synthesized in
the 5' → 3' by
DNA polymerase
78
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Lagging Strand t2
FILTER

The RNA primers are

then removed and new
deoxyribonucleotides
are added to the gaps,
hig

h2

where the RNA was

present
79
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Lagging Strand t2
FILTER

DNA Polymerase
continues with the
synthesis of the new
hig

h2

DNA strand

80
t3
t1
 Lesson 2

Mechanism of DNA Replication

Synthesis of the Lagging Strand t2
FILTER

Finally, DNA ligase

(an enzyme) joins the
hig
deoxyribonucleotides
together, thus
h2

completing the
lagging strand
81
t3
t1
 Lesson 2

Mechanism of DNA Replication

Termination of Replication t2
FILTER

In eukaryotes,
termination of
replication is
hig

h2

poorly understood

82
t3
t1
 Lesson 2

Mechanism of DNA Replication

Termination of Replication t2
FILTER

Eukaryotes have
linear DNA, and
therefore use
hig

telomeres, which are h2

GT rich repeating
units that 'protect'
the end of the DNA
83
t3
t1
 Featured image
FILTER

Module 2 Module Lessons

DNA Replication

Dashboard

Introduction
t1 t2 t3

Lessons
hig

Lesson 1 h1
Lesson 2 title
h2
Lesson 3 h3

Assignments
Method of Mechanism of Rate of
Replication Replication Replication

Start Start Start

84
 Module 2

DNA Replication

t3
h3

hig

Lesson 3

Rate of Replication t2

Find out how long it takes for DNA to replicate

t1

Learn Now
 Lesson 3

Rate of Replication
t3
FILTER

The single molecule

of DNA, that is the
hig

E. coli genome h3

contains 4.7 x 106

nucleotide pairs

86
t2
t1
 Lesson 3

Rate of Replication
t3
FILTER

DNA replication
begins at a single,
fixed location in this
molecule, called the
hig
replication origin,
proceeds at about
h3

1000 nucleotides per

second, and thus is
done in no more
than 40 minutes
87
t2
t1
 Lesson 3

Rate of Replication
t3
FILTER

Due to the precision

of the process (which
includes a "proof-
reading" function),
hig

the job is done with h3

only about one

incorrect nucleotide
for every 109
nucleotides inserted 88
t2
t1
 Lesson 3

Rate of Replication
t3
FILTER

In other words,
more often than
hig

not, the E. coli h3

genome (4.7 x 106)

is copied without
error!
89
t2
t1
 Lesson 3

Rate of Replication
t3
FILTER

The Genome of
complex
hig

eukaryotes is huge h3

as compared to
procaryotes

90
t2
t1
 Lesson 3

Rate of Replication
t3
FILTER

The speed of DNA

replication for
hig

human is about h3

50-75 nucleotides
per second per
replication fork
91
t2
t1
 Lesson 3

Rate of Replication
t3
FILTER

However, the
genome can be
copied only in a
few hours because
many replication
hig

h3

forks take place at

the same time
(multiple initiation
sites) 92
t2
t1
 Featured image
FILTER

Module 2 Module Lessons

DNA Replication

Dashboard

Introduction
t1 t2 t3

Lessons
hig

Lesson 1 h1
Lesson 2 title
h2
Lesson 3 h3

Assignments
Method of Mechanism of Rate of
Replication Replication Replication

Start Start Start

93
 Module 2

DNA Replication

hig

t3

End of Module 2 t2

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