13

DNA and Its Role in Heredity

Adapted from “Life: The Science of Biology” 12th edition by Sadava, Hillis, Heller, Hacker.

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Chapter Outline

What is the evidence that the gene is DNA?

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The evidence that the gene is DNA

By 1920, it was known that chromosomes consisted of DNA and proteins.

A new dye was developed to stain DNA, which provided evidence that DNA
was the genetic material:
• It was in the right place- in the nucleus and chromosomes.
• It varied among species- different amount in different species.
• It was present in the right amount- twice in somatic cells vs gametes.

Experiments on bacteria and viruses demonstrated DNA is the genetic

material.

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The evidence that the gene is DNA

Frederick Griffith: worked with two strains of Streptococcus pneumonia:

- S strain: Virulent. Smooth colonies with polysaccharide capsule.
S strain was virulent because it was protected from the immune
system by the capsule.
- R strain: Non-virulent. Rough look colonies. No Capsule.

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Figure 13.1 Genetic Transformation (Experiment)

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 The evidence that the gene is DNA

 Some cells of R were transformed into S and reproduced- genetic change.

 A chemical from dead cells of one strain produced a heritable change in the other
strain. The transforming principle.
 Conclusion: Griffith's experiments showed that a molecule in a lethal type of bacteria
can transform nonvirulent bacteria into virulent.

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The evidence that the gene is DNA

The chemical transforming substance was identified by Avery et al.:

 Treated samples with enzymes that destroy different molecules: proteins, RNA, DNA.
 Results: - after proteins and RNA destruction, R strain still transformed into S.
- after destroying DNA, the transforming activity was lost.

- Then, they injected DNA alone and

caused bacterial transformation.

Conclusion:
The DNA is responsible for
producing the polysaccharide
capsule was transferred into the R
cells.

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 Summary

 Griffith’s experiments demonstrated a transforming principle causes

heritable changes in other cells.
 The location and quantity of DNA in a cell suggested it may be DNA.
 The Hershey-Chase experiment established conclusively that it was
DNA and not protein.
 Transfection is the genetic transformation of eukaryotic cells.

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What is the structure of DNA?
 The structure of DNA was determined using many lines of evidence.
 One crucial piece came from X-ray crystallography.
 A purified substance can be made to form crystals; position of atoms is inferred
by the pattern of diffraction of X rays passed through the crystallized substance.
Rosalind Franklin:
 Prepared crystallographs from DNA fibers.
 Her images suggested a spiral model.

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What is the structure of DNA?

Chemical composition of DNA also provided clues:

 DNA is a polymer of nucleotides: each containing
Deoxyribose, a phosphate group, and a nitrogen-
containing base.

In 1950, Erwin Chargaff found in the DNA from many

different species that:
Amount of A = amount of T
Amount of C = amount of G

* the amount of purines = the amount of pyrimidines

Chargaff’s rule.

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What is the structure of DNA?

Francis Crick and James Watson

used model building and combined all the
knowledge of DNA to determine its structure.

• Franklin’s X-ray crystallography

* the molecule is double stranded and helical.

• Chargaff rule on paired bases suggested

antiparallel strands.

• In 1953, Watson and Crick established the

general structure of DNA.

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What is the structure of DNA?

Complementary base pairing:

 Adenine pairs with thymine by 2 H-bonds.
 Cytosine pairs with guanine by 3 H-bonds.
 Every base pair consists of one purine and
one pyrimidine.

Antiparallel strands: Direction of the strand is

determined by the sugar–phosphate bonds.
 Phosphate groups connect to the 3′ C of
one sugar, and the 5′ C of the next sugar.
 The two chain ends differ:
- a free 5′ phosphate group—the 5′ end.
- a free 3′ hydroxyl group (OH)—the 3′ end.

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Concept 13.2 DNA Has a Structure That Suits Its Function (6)

The X-ray diffraction data indicated that

• The bases are on the inside of each strand.
• The sugar-phosphate groups are on the outside of each
strand.
• The chains run in opposite directions—antiparallel.
Key features of DNA structure:
• It is a double-stranded helix
• It is right-handed helix
• It is antiparallel
• The strands are held together by complementary base pairing
• The outer edges of the bases are exposed in major and minor
grooves

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Figure 13.8 Base Pairs in DNA Can Interact with Other Molecules

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Concept 13.2 DNA Has a Structure That Suits Its Function (11)

The double-helix structure is essential to DNA function:

• With millions of nucleotides, the base sequences store a
huge amount of genetic information.
• Susceptible to mutations, simple changes in the linear
sequence of base pairs.
• Precise replication in cell division is possible by
complementary base pairing.
• Genetic information is expressed as the phenotype—
nucleotide sequences determine sequences of amino
acids in proteins; proteins determine phenotypes

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 Summary

 Chargaff’s rule states that the amount of adenine in DNA equals the
amount of thymine, the amount of guanine equals the amount of cytosine.

 X-ray crystallography showed that DNA is helical. Watson and Crick

proposed that DNA is a double stranded helix in which strands are
antiparallel.

 Complementary base pairing holds the bases together by hydrogen

bonding.

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Chapter Outline

13.3 How is DNA replicated?

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Concept 13.3 DNA Is Replicated Semiconservatively (1)

Researchers found that DNA could be replicated in

a test tube using
• Nucleoside monomers of DNA
• DNA molecules to serve as templates
• DNA polymerase
• Salts and pH buffer
This confirmed that DNA contains the information
needed for its own replication.

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Concept 13.3 DNA Is Replicated Semiconservatively (2)

Possible replication patterns:

• Semiconservative: each parent strand is a
template; new molecules have one old and one
new strand
• Conservative: original molecule serves as a
template only
• Dispersive: DNA fragments are templates; old
and new pieces are assembled into new
molecules

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Figure 13.9 Three Models for DNA Replication

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 How is DNA replicated?

DNA replication is semiconservative:

* the two new DNA molecules have one old and one new strand.

Meselson and Stahl:

Density labeling – “The most beautiful experiment in biology”.
 Escherichia coli growing in media with heavy isotope 15N, and14N. Combined
extracts and found two separated bands of DNA.
 E coli from 15N was passed to 14N. Then collected DNA at time intervals (20 min).
F1 = All DNA intermediate density
F2 = DNA 2 bands: intermediate and light

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 How is DNA replicated?
DNA replication occurs in two steps:
1. The double helix is unwound (2 template strands for base pairing).
2. New nucleotides are added to the new strands (at the 3′ end).

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Concept 13.3 DNA Is Replicated Semiconservatively
(7)

DNA replication starts when a large protein complex (pre-

replication complex) binds to a region called origin of
replication (ori).

In E. coli, DNA is unwound and replication proceeds in both

directions, forming two replication forks.

Eukaryote chromosomes are much longer and have multiple

origins of replication.

Otherwise, it would take weeks to replicate chromosomes,

which have up to a billion base pairs.

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 Figure 13.11 The Origin of DNA Replication

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 How is DNA replicated?

Before replication, DNA helicase unwind the DNA at ori (DNA sequence in the
chromosome where replication starts).

DNA replication begins with a short primer

synthesized by the enzyme Primase.

The primer is a starter strand of RNA,

complementary to the DNA template.

After the primer, DNA polymerase binds to add

nucleotides to the 3′ end of new strand.

Single-strand binding proteins- keep strands

from getting back together.

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 How is DNA replicated?

The site where DNA unwinds opens DNA like a zipper, in 1 direction. So:
- One new “leading strand”, will grow from original DNA 3′ end to the fork (5’ end).
Can it be the same with the other strand?
- One new “lagging strand” will grow in small, discontinuous stretches from the
fork to it’s 5’ end — Okazaki fragments.

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How is DNA replicated?

Each Okazaki fragment builds its own primer.

DNA polymerase adds nucleotides until reaching the primer of the previous
fragment.

DNA ligase replace the primer to DNA and fill in the gaps between segments.

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Concept 13.3 DNA Is Replicated Semiconservatively (17)

Eukaryote chromosomes have repetitive

sequences at the ends called telomeres.
In humans, the sequence is TTAGGG-3′, repeated
about 2,500 times.
The repeats bind proteins that prevent the DNA
repair system from recognizing chromosome
ends as breaks.

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Concept 13.3 DNA Is Replicated Semiconservatively (18)

On lagging strands, when the terminal Okazaki

primer is removed, no DNA can be synthesized
to replace it (no 3′ end).
The short piece of single stranded DNA is
removed, and the chromosome becomes shorter
with each replication.
After many divisions, genes may be lost and the
cell dies.

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Figure 13.18 Telomeres and Telomerase

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 SUMMARY

 DNA replication is semiconservative, each parent strand acts as a template

strand for the synthesis of a new strand.

 DNA polymerase catalyses the addition of nucleotides to the 3’ end of each

strand by complementary base pairing.

 The leading strand is synthesized continuously while the lagging strand in

Okazaki fragments, which are then joined together by DNA ligase.

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Chapter Outline

13.4 How are errors in DNA repaired?

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Concept 13.4 Errors in DNA Can Be Repaired (1)

DNA polymerases make mistakes and DNA can be

damaged by chemicals, UV radiation, and other
threats.
Cells have three repair mechanisms:
1. Proofreading: DNA polymerase recognizes
mismatched pairs and removes incorrectly
paired bases. Catches 99% or more
mismatches.

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Concept 13.4 Errors in DNA Can Be Repaired (2)

2. Mismatch repair: Newly replicated DNA is

scanned for mistakes by other proteins and
mismatches can be corrected.

3. Excision repair: Enzymes scan DNA for

damaged bases—they are excised and DNA
polymerase I adds the correct ones.

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Figure 13.19 DNA Repair Mechanisms

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 SUMMARY

 DNA polymerase makes errors during replication.

 DNA is also subject to natural changes and chemical damage.

 DNA can be repaired by three different mechanisms:

proofreading, mismatch repair, and excision repair.

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Chapter Outline

13.5 How does the polymerase chain

reaction amplify DNA?

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Concept 13.5 The Polymerase Chain Reaction Amplifies DNA (1)

The principles of DNA replication were used to

develop the polymerase chain reaction (PCR)
technique.
An automated process makes multiple copies of
short DNA sequences for genetic manipulation
and research (DNA amplification).

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Concept 13.5 The Polymerase Chain Reaction Amplifies DNA (2)

A PCR mixture contains:

• A sample of double-stranded DNA (the
template)
• Two artificially synthesized primers
• Four dNTPs
• DNA polymerase that can tolerate high
temperatures
• Salts and pH buffer

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Concept 13.5 The Polymerase Chain Reaction Amplifies DNA (3)

In PCR amplification:
• DNA strands are separated (denatured) by
heating
• Reaction is cooled to allow primers to bind
(anneal) to template strands
• Reaction is warmed; DNA polymerase catalyzes
new strands
• The sequence is repeated many times

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Figure 13.20 The Polymerase Chain Reaction

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Concept 13.5 The Polymerase Chain Reaction Amplifies DNA (4)

Base sequences at the 3ʹ ends of the DNA strands

must be known, so that primers can be made.
The specificity of the primers is a key to the power
of PCR.
The DNA polymerase that does not denature at
high temperatures (90°C) was taken from a hot
springs bacterium, Thermus aquaticus.

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 SUMMARY

 Polymerase chain reaction makes use of the knowledge from DNA

replication to make multiple copies of DNA sequences.

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