Download Chapter 10 - McGraw Hill Higher Education

Essentials of
The Living World
First Edition
GEORGE B. JOHNSON
10
The New Biology
PowerPoint® Lectures prepared by Johnny El-Rady
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10.1 Genomics
The full complement of genetic information of an
organism is its genome
Genomics is a new field of biology concerned with
the sequencing and study of genomes
The first genome to be sequenced was that of the
virus FX174
Frederick Sanger in 1977 obtained the sequence
of this 5,375 genome
The advent of automatic DNA sequencing machines
has facilitated the sequencing of larger genomes
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Sequencing DNA
DNA is first amplified
The DNA fragments are then mixed with
Primers + DNA polymerase
A supply of the four nucleotides
A smaller supply of chemically-tagged nucleotides
that terminate replication
The DNA is denatured into single-strands allowing
DNA replication to proceed
The addition of a chemically-tagged nucleotide to the
growing chain halts DNA replication
Thus the mixture will contain double-stranded
DNA of various lengths
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Sequencing DNA
The DNA mixture is separated by fragment size
using gel electrophoresis
Examination of the fragments from shortest to
longest reveals the nucleotide sequence of the DNA
Linking the sequence of the various DNA fragments
will yield the sequence of the entire genome
The scanning and analysis of the gel is greatly
facilitated by the use of computers
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Fig. 10.1 How to sequence DNA
One color
corresponds to
each nucleotide
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TABLE 10.1 EUKARYOTIC GENOMES
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TABLE 10.1 EUKARYOTIC GENOMES
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10.2 The Human Genome
The sequence of the entire human genome was
reported on June 26, 2000
It consists of 3.2 billion base pairs
If the human genome were a book
It would be 500,000 pages long
It would take about 60 years to read at the rate of
8 hours a day, every day, at five bases a second
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Geography of the Genome
The number of genes in humans is only about
25,000-30,000
However, there are about 4 times more mRNA
molecules
The genes are divided into exons and introns
Thus alternative mRNA splicing can generate
much more mRNA than there are genes
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Fig. 10.2
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Geography of the Genome
Genes are not distributed evenly throughout the
human genome
Chromosome 19 is small yet packed with genes
Chromosomes 4 and 8 are large yet have few
genes
On most chromosomes, clusters rich in genes are
scattered between vast stretches of “barren” DNA
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DNA That Codes for Proteins
The human genome contains four different classes
of protein-encoding genes
1. Single-copy genes
Most genes fit in this class
Silent copies, inactivated by mutation, are
called pseudogenes
2. Segmental duplications
Blocks of similar genes in the same order are
found throughout the human genome
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DNA That Codes for Proteins
The human genome contains four different classes
of protein-encoding genes
3. Multigene families
Groups of related but distinctly different genes
that often occur together in cluster
Arose from a single ancestral sequence
4. Tandem clusters
DNA sequences repeated thousands of times
in tandem array
The rRNA genes, for example
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Noncoding DNA
Only 1-1.5% of the human genome is coding DNA
There are four major types of noncoding DNA
1. Noncoding DNA within genes
Together introns make up about 24% of the
human genome
2. Structural DNA
~ 20% of the genome is constitutive
heterochromatin
Located near centromeres and telomeres
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Noncoding DNA
Only 1-1.5% of the human genome is coding DNA
There are four major types of noncoding DNA
3. Repeated sequences
Simple sequence repeats (SSRs)
Two- or three-nucleotide sequences repeated
thousands of times
Constitute ~ 3% of the human genome
Duplicated Sequences
Repeated sequences, other than SSRs
Constitute ~ 7% of the human genome
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Noncoding DNA
Only 1-1.5% of the human genome is coding DNA
There are four major types of noncoding DNA
4. Transposable elements
Make up ~ 45% of the human genome
They include
LINEs
Long interspersed elements
~ 6,000 DNA bases long
Active transposons
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Noncoding DNA
Only 1-1.5% of the human genome is coding DNA
There are four major types of noncoding DNA
4. Transposable elements
Make up ~45% of the human genome
They include
Alu sequences
~ 300 DNA bases long
Have no transposition machinery
Reside within, and transpose with, LINEs
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10.3 A Scientific Revolution
Genetic engineering is the process of moving genes
from one organism to another
Having a major impact on agriculture & medicine
Fig. 10.3
Producing insulin
Curing disease
Increasing yields
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Restriction Enzymes
Restriction enzymes bind to specific short sequences
(usually 4- to 6- bases long) on the DNA
The nucleotide sequence on both
DNA strands is identical when
read in opposite directions
GAATTC
CTTAAG
Most restriction enzymes cut the DNA in a
staggered fashion
This generates “sticky” ends
These ends can pair with any other DNA
fragment generated by the same enzyme
The pairing is aided by DNA ligase
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Fig. 10.4
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Stages of a Genetic Engineering Experiment
All gene transfer experiments share four distinct
stages
1.
2.
3.
4.
Cleaving DNA
Producing recombinant DNA
Cloning
Screening
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Fig. 10.5
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10.4 Genetic Engineering
and Medicine
Genetic engineering has been used in many medical
applications
1. Production of proteins to treat illnesses
2. Creation of vaccines to combat infections
3. Replacement of defective genes
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Making “Magic Bullets”
In diabetes, the body is unable to control levels of
sugar in the blood because of lack of insulin
Diabetes can be cured if the body is supplied
with insulin
The gene
encoding insulin
has been
introduced into
bacteria
Fig. 10.3
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Making “Magic Bullets”
Other genetically engineered drugs include
Has only one
extra gene:
HGH
Anticoagulants
Used to treat heart
attack patients
Factor VIII
Used to treat
hemophilia
Human growth
hormone (HGH)
Used to treat
dwarfism
Fig. 10.6
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Piggyback Vaccines
Genetic engineering has also been used to create
subunit vaccines against viruses
A gene encoding a viral protein is put into the DNA
of a harmless virus and injected into the body
The viral protein will elicit antibody production in
the animal
A novel kind of vaccine was introduced in 1995
The DNA vaccine uses plasmid vectors
It elicits a cellular immune response,
rather than antibody production
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Fig. 10.7 Constructing a piggyback vaccine for the herpes simplex virus
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10.5 Genetic Engineering
and Agriculture
Successful manipulation of the genes of crop plants
has improved the quality of these plants
Pest resistance
Leads to a reduction in the use of pesticides
Bt, a protein produced by soil bacteria, is harmful
to pests but not to humans
The Bt gene has been introduced into tomato
plants, among others
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10.5 Genetic Engineering
and Agriculture
Glyphosateresistant plants
Herbicide resistance
Crop plants have
been created that
are resistant to
glyphosate
Fig. 10.9
Glyphosatesensitive plants
Petunias
Herbicide resistance offers two main advantages
1. Lowers the cost of producing crops
2. Reduces plowing and conserves the top soil
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10.5 Genetic Engineering
and Agriculture
More Nutritious Crops
Worldwide, two major deficiencies are iron and
vitamin A
Deficiencies are especially severe in developing
countries where the major staple food is rice
Ingo Potrykus, a Swiss
bioengineer, developed
transgenic “golden” rice
to solve this problem
Fig. 10.10
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Fig. 10.10 Transgenic “golden” rice
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Potential Risks of Genetically Modified (GM) Crops
The promise of genetic engineering is very much in
evidence
However, it has generated considerable
controversy and protest
Are genetic engineers “playing God” by
tampering with the genetic material?
Two sets of risks need to be considered
1. Are GM foods safe to eat?
2. Are GM foods safe for the environment?
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Potential Risks of Genetically Modified (GM) Crops
1. Are GM foods safe to eat?
The herbicide glyphosate blocks the synthesis of
aromatic amino acids
Humans don’t make any aromatic amino acids,
so glyphosate doesn’t hurt us
However, gene modifications that render plants
resistant to glyphosate may introduce novel
proteins
Moreover, introduced proteins may cause
allergies in humans
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Potential Risks of Genetically Modified (GM) Crops
2. Are GM foods safe for the environment?
Three legitimate concerns are raised
1. Harm to other organisms
Will other organisms be harmed
unintentionally?
2. Resistance
Will pests become resistant to pesticides?
3. Gene flow
What if introduced genes will pass from GM
crops to their wild or weedy relatives?
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10.6 Reproductive Cloning
In 1938, the German embryologist Hans Spemann
proposed what he called a “fantastical experiment”:
Replace the nucleus of an egg cell with the
nucleus of another cell
Early attempts to clone animals in this way failed
The breakthrough was the following insight
Starvation will synchronize cells at the same
point in the cell cycle
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Wilmut’s Lamb
Reproductive biologist Ian Wilmut and his colleagues
were able to clone the first animal in 1997
Mammary cells were removed from the udder of a
six-year old sheep
The nucleus was removed from an egg cell taken
from another sheep
Both cells were synchronized to a resting state
The nucleus from the mammary cell was
transferred to the enucleated egg cell
An electric shock was applied to start cell division
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Wilmut’s Lamb
The successful embryos (about 30 in 277 tries)
were transplanted into surrogate mother sheep
On July 5, 1996, “Dolly” was born
Only 1 of 277 tries succeeded
However, Wilmut proved that reproductive
cloning is possible
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Fig. 10.11 Wilmut’s animal cloning experiment
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Problems with Reproductive Cloning
Since Dolly, scientists have successfully cloned
sheep, mice, cattle, goats and pigs
However, problems and complications arise,
leading to premature death
Dolly died in 2002, having lived only half a
normal sheep life span
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The Importance of Genomic Imprinting
During gamete development, the DNA undergoes a
process termed genomic imprinting
The process involves the methylation (addition of
–CH3 groups) to cytosine residues in the DNA
This locks genes in either the “on” or “off”
position
Normal animal development requires chemical
reprogramming of the DNA
Takes months to years in adult reproductive
tissues
Cloning fails because there is not enough time for
the re-programming to be done properly
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10.7 Embryonic Stem Cells
The blastocyst, an early embryo, consists of
A protective outer layer that will form the placenta
Inner cell mass that will form the embryo
The inner cell mass consists of embryonic stem cells
These are pluripotent
Capable of forming the entire organism
As development proceeds, cells lose their
pluripotency
They become committed to one type of tissue
They are then called adult stem cells
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10.7 Embryonic Stem Cells
Embryonic stem cells could be used to restore
tissues lost or damaged due to accident or disease
Experiments have already been tried successfully
in mice
Damaged spinal neurons have been partially
repaired
The course of development is broadly similar in all
mammals
Therefore, the experiments in mice are very
promising
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Fig. 10.12
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10.7 Embryonic Stem Cells
The research in human
embryonic stem cells is
associated with two
serious problems
1. Finding a source
Harvesting them from
discarded embryos
raises ethical issues
Fig. 10.13 Human embryonic stem
cells
2. Immunological rejection
Implanted stem cells will likely be rejected by
the immune system of the individual
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10.7 Embryonic Stem Cells
Therapeutic cloning follows this basic approach:
A cell is obtained from an individual who lost a
tissue function due to an accident or disease
It is cloned to produce an embryo
Embryonic stem cells are harvested and grown
in tissue culture
The stem cells are then injected back into the
same individual
There, they divide and ultimately differentiate
into healthy tissue
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Fig. 10.15 How human embryos might be used for therapeutic cloning
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Fig. 10.15 How human embryos might be used for therapeutic cloning
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10.7 Embryonic Stem Cells
Therapeutic cloning solves the problem of immune
rejection
Cells are cloned from the individual’s own tissues,
Therefore, they pass the immune system’s
“self” identity check
However, the process is still controversial
Some fear that the cloned embryo might be
brought to term by inserting it into a human uterus
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10.8 Gene Therapy
Gene therapy involves the introduction of “healthy”
genes into cells that lack them
It was first used successfully in 1990
Two girls were cured of a rare blood disorder
caused by a defective adenosine deaminase gene
The girls stayed healthy
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10.8 Gene Therapy
Researchers then set out to apply gene therapy to
cystic fibrosis
In 1994, the technique was first tried on mice
A normal copy of the gene, cf, was added to the vector
adenovirus
The virus was then squirted into the lungs of mice that
carried a defective cf gene
The mice had their immune systems disabled
The “healthy” gene was thus introduced into lung cells
And the mice were successfully cured!
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10.8 Gene Therapy
Researchers then targeted humans in 1995
The same basic approach was used as was used
with mice
For eight weeks, the gene therapy seemed
successful
However, the gene modified-cells in the patients’
lungs came under attack by the immune system
The healthy genes were lost, and with them the
chances for a cure
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A comprehensive 1995 review of human gene
therapy trials revealed three problems
1. The adenovirus elicits a strong immune
response
It causes the common cold, so antibodies were formed
due to previous colds
2. In rare cases, the immune response can be
very severe
If many patients are treated, a few may die
3. The adenovirus inserts its DNA randomly in
human chromosomes
This will cause mutations and potentially cancers
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More Promising Vectors
Within a few years, researchers had a much more
promising vector
A tiny virus called adeno-associated virus (AAV)
AAV only has two genes and thus needs
adenovirus to replicate
AAV has several advantages over adenovirus
1. It inserts genes into human DNA less frequently
2. It does not elicit a strong immune response
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Success with the AAV Vector
In 1999, AAV successfully cured anemia in rhesus
monkeys
AAV was also used to cure dogs of a hereditary
disorder leading to retinal degeneration & blindness
And human trials are under way again!
In 2000, scientists performed the first gene
therapy experiment for muscular dystrophy
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Fig. 10.16 Using gene therapy to cure a retinal degenerative disease in dogs
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