Ch12 notes

Chapter 12

DNA Technology and Genomics

PowerPoint Lectures for

Campbell Biology: Concepts & Connections, Seventh Edition
Reece, Taylor, Simon, and Dickey
© 2012 Pearson Education, Inc.

Lecture by Edward J. Zalisko

Figure 12.0_1

Chapter 12: Big Ideas

Gene Cloning

Genetically Modified
Organisms

DNA Profiling

Genomics

What are clones?
Cells or individuals derived from a single ancestor.
The idea of cloning is not new.
– Animal cloning has been done since the late 1800s.

Identical twins are natural clones

Types of Cloning

 Reproductive Cloning
– Used to produce a new individual that has identical
DNA as an already living individual

 Therapeutic Cloning
– Used to produce stem cells to study human
development and cure disease.
– Growing Body Parts – 60 Minutes video
– Replacing Body Parts - NOVA

Animal Cloning
 First animal clone was a sea urchin by Hans
Dreisch in the late 1800s.
 Methods of Cloning
– Embryo splitting
– When an embryo reaches 8-16 cells they are split
– Each will develop into genetically identical embryos

– Nuclear transfer
– Nucleus is removed from an egg cell
– Egg cell is fused with cells from the individual to be cloned
– The egg cells are then transplanted into a surrogate
mother for development.

 Dolly was the first successful clone produced by nuclear
transfer of an adult cell.
– 1997

 Animal cloning has had a great impact on farming.
– We can produce herds of animals with desirable traits (superior
wool, milk or meat production)

• Problem with animal cloning in
agriculture:
• Inefficient
• Stigma attached to “cloned
food/drink”
Wired Science – “Got Clones”

 Cloning extinct animals
– Video – Frozen Zoo – 60 Minutes

– Tasmanian Tiger
– Video – Bringing Back the Tasmanian Tiger (Learn 360)

– Wooly Mammoth
– Video – Resurrecting the Extinct – 60 Minutes
– Video – Waking the Baby Mammoth – National Geographic

12.1 Genes can be cloned in recombinant plasmids
 Biotechnology is the manipulation of organisms
or their components to make useful products.
 DNA technology is the set of modern techniques
used to study and manipulate genetic material.


 Genetic engineering involves manipulating
genes for practical purposes.
– Gene cloning leads to the production of multiple,
identical copies of a gene-carrying piece of DNA.
– Recombinant DNA is formed by joining nucleotide
sequences from two different sources.
– One source contains the gene that will be cloned.
– Another source is a gene carrier, called a vector.
– Plasmids (small, circular DNA molecules independent of the
bacterial chromosome) are often used as vectors.


 Steps in cloning a gene
1. Plasmid DNA is isolated.
2. DNA containing the gene of interest is isolated.
3. Plasmid DNA is treated with a restriction enzyme that
cuts in one place, opening the circle.
4. DNA with the target gene is treated with the same
enzyme and many fragments are produced.
5. Plasmid and target DNA are mixed and associate with
each other.


6. Recombinant DNA molecules are produced when
DNA ligase joins plasmid and target segments
together.
7. The recombinant plasmid containing the target gene
is taken up by a bacterial cell.
8. The bacterial cell reproduces to form a clone, a group
of genetically identical cells descended from a single
ancestral cell.

Animation: Cloning a Gene

Figure 12.1B

E. coli bacterium
Plasmid

Bacterial
chromosome

1

A cell with DNA
containing the gene
of interest
2

A plasmid
is isolated.

The cell’s DNA
is isolated.

Gene of
interest
3

DNA

The plasmid is cut
with an enzyme.

Examples of gene use
4

The cell’s DNA is cut
with the same enzyme.

Gene
of interest

5

6

The targeted fragment
and plasmid DNA
are combined.

DNA ligase is added,
which joins the two
DNA molecules.

Examples of protein use

Recombinant
DNA
plasmid

Gene
of interest
7

The recombinant plasmid
is taken up by a bacterium
through transformation.

Recombinant
bacterium
8

Clone
of cells

Genes may be inserted
into other organisms.

The bacterium
reproduces.

9

Harvested
proteins
may be
used
directly.

12.2 Enzymes are used to “cut and paste” DNA
 Restriction enzymes cut DNA at specific
sequences.
– Each enzyme binds to DNA at a different restriction
site.
– Many restriction enzymes make staggered cuts that
produce restriction fragments with single-stranded
ends called “sticky ends.”
– Fragments with complementary sticky ends can
associate with each other, forming recombinant DNA.

 DNA ligase joins DNA fragments together.
Animation: Restriction Enzymes

Figure 12.2_s4
1

DNA
Restriction enzyme
recognition sequence

A restriction
enzyme cuts
the DNA into
fragments.
2

Sticky
end
A DNA fragment
from another
source is added.

Restriction
enzyme

Sticky
end

Gene of
interest

3

Two (or more)
fragments stick
together by
base pairing.
4

DNA ligase
pastes the
strands together.

5

DNA ligase

Recombinant
DNA molecule

12.5 Nucleic acid probes identify clones carrying
specific genes
 Nucleic acid probes bind very selectively to
cloned DNA.
– Probes can be DNA or RNA sequences
complementary to a portion of the gene of interest.
– A probe binds to a gene of interest by base pairing.
– Probes are labeled with a radioactive isotope or
fluorescent tag for detection.


Figure 12.5

Radioactive
nucleic acid probe
(single-stranded DNA)

The probe is mixed with
single-stranded DNA
from a genomic library.

Single-stranded
DNA

Base pairing
highlights the
gene of interest.

12.6 Recombinant cells and organisms can
mass-produce gene products
 Recombinant cells and organisms constructed by
DNA technologies are used to manufacture many
useful products, chiefly proteins.
 Bacteria, yeasts, and mammalian cells

 Bacteria are often the best organisms for
manufacturing a protein product because bacteria
– have plasmids and phages available for use as genecloning vectors,
– can be grown rapidly and cheaply,
– can be engineered to produce large amounts of a
particular protein, and
– often secrete the proteins directly into their growth
medium.

12.6 Recombinant cells and organisms can
mass-produce gene products
 Pharmaceutical researchers are currently exploring
the mass production of gene products by
– whole animals or plants.

 Recombinant animals
– are difficult and costly to produce
– must be cloned to produce more animals with the same
traits.


12.7 DNA technology has changed the
pharmaceutical industry and medicine
 Products of DNA technology are already in use.
– Therapeutic hormones produced by DNA technology
include
– insulin to treat diabetes
– human growth hormone to treat dwarfism.

– DNA technology is used to
– test for inherited diseases,
– detect infectious agents such as HIV, and
– produce vaccines, harmless variants (mutants) or derivatives
of a pathogen that stimulate the immune system.


12.8 Genetically modified organisms are
transforming agriculture
 Genetically modified (GM) organisms contain
one or more genes introduced by artificial means.
 Transgenic organisms contain at least one gene
from another species.
 The most common vector used to introduce new
genes into plant cells is
– a plasmid from a soil bacterium called the Ti plasmid.


Figure 12.8A_s3

Agrobacterium
tumefaciens
DNA containing the
gene for a desired trait

Ti
plasmid

Restriction
site

1

The gene is
inserted into
the plasmid.

Recombinant
Ti plasmid

Plant cell

2

The recombinant
plasmid is
introduced into
a plant cell.
DNA carrying
the new gene
3

The plant cell
grows into
a plant.
A plant
with the
new trait

12.8 Genetically modified organisms are
transforming agriculture
 GM plants are being produced that
– are more resistant to herbicides and pests
– provide nutrients that help address malnutrition.

 GM animals are being produced with improved
nutritional or other qualities.


12.9 Genetically modified organisms raise
concerns about human and environmental
health
 Concerns related to GM organisms include the
potential
– introduction of allergens into the food supply and
– spread of genes to closely related organisms.

 Regulatory agencies are trying to address the
– safety of GM products,
– labeling of GM produced foods, and
– safe use of biotechnology.


12.10 Gene therapy may someday help treat a
variety of diseases
 Gene therapy aims to treat a disease by
supplying a functional allele.
 One possible procedure is the following:
1. Clone the functional allele and insert it in a retroviral
vector.
2. Use the virus to deliver the gene to an affected cell
type from the patient, such as a bone marrow cell.
3. Viral DNA and the functional allele will insert into the
patient’s chromosome.
4. Return the cells to the patient for growth and division.

12.10 CONNECTION: Gene therapy may
someday help treat a variety of diseases
 Gene therapy is an
– alteration of an afflicted individual’s genes and
– attempt to treat disease.

 Gene therapy may be best used to treat disorders
traceable to a single defective gene.


Figure 12.10

Cloned gene
(normal allele)

1 An RNA version of

a normal human
gene is inserted
into a retrovirus.

RNA genome of virus
Retrovirus

2 Bone marrow cells

are infected with
the virus.

3 Viral DNA carrying the

human gene inserts into
the cell’s chromosome.
Bone marrow
cell from the patient
4 The engineered

cells are injected
into the patient.

Bone
marrow

 The first successful human gene therapy trial in
2000
– tried to treat ten children with SCID (severe combined
immune deficiency),
– helped nine of these patients, but
– caused leukemia in three of the patients, and
– resulted in one death.


 The use of gene therapy raises many questions.
– How can we build in gene control mechanisms that
make appropriate amounts of the product at the right
time and place?
– How can gene insertion be performed without harming
other cell functions?
– Will gene therapy lead to efforts to control the genetic
makeup of human populations?
– Should we try to eliminate genetic defects in our
children and descendants when genetic variety is a
necessary ingredient for the survival of a species?

12.11 The analysis of genetic markers can
produce a DNA profile
 DNA profiling is the analysis of DNA fragments to
determine whether they come from the same
individual.


12.12 The PCR method is used to amplify DNA
sequences
 Polymerase chain reaction (PCR) is a method of
amplifying a specific segment of a DNA molecule.
 PCR
– is a three-step cycle that
– doubles the amount of DNA in each turn of the cycle.


12.12 The PCR method is used to amplify DNA
sequences
 The advantages of PCR include
– the ability to amplify DNA from a small sample,
– obtaining results rapidly, and
– a reaction that is highly sensitive, copying only the
target sequence.


12.13 Gel electrophoresis sorts DNA molecules
by size
 Gel electrophoresis can be used to separate
DNA molecules based on size as follows:
1. A DNA sample is placed at one end of a porous gel.
2. Current is applied and DNA molecules move from the
negative electrode toward the positive electrode.
3. Shorter DNA fragments move through the gel matrix
more quickly and travel farther through the gel.
4. DNA fragments appear as bands, visualized through
staining or detecting radioactivity or fluorescence.
5. Each band is a collection of DNA molecules of the
same length.
Video: Biotechnology Lab


Figure 12.13

A mixture of DNA
fragments of
different sizes
Longer
(slower)
molecules

Power
source
Gel
Completed
gel

Shorter
(faster)
molecules

12.14 STR analysis is commonly used for DNA
profiling
 Repetitive DNA consists of nucleotide sequences
that are present in multiple copies in the genome.
 Short tandem repeats (STRs) are short nucleotide
sequences that are repeated in tandem,
 STR analysis
– compares the lengths of STR sequences at specific sites
in the genome and
– typically analyzes 13 different STR sites.


Figure 12.14A

STR site 1

STR site 2

Crime scene
DNA
The number of short
tandem repeats match

Suspect’s
DNA

The number of short tandem
repeats do not match

Figure 12.14B

Crime
scene
DNA

Suspect’s
DNA

Longer STR fragments

Shorter STR fragments

12.15 CONNECTION: DNA profiling has provided
evidence in many forensic investigations
 DNA profiling is used to
– determine guilt or innocence in a crime,
– settle questions of paternity,
– identify victims of accidents, and
– probe the origin of nonhuman materials.


12.16 RFLPs can be used to detect differences in
DNA sequences
 A single nucleotide polymorphism (SNP) is a
variation at a single base pair within a genome.
 Restriction fragment length polymorphism
(RFLP) is a change in the length of restriction
fragments due to a SNP that alters a restriction
site.
 RFLP analysis involves
– producing DNA fragments by restriction enzymes and
– sorting these fragments by gel electrophoresis.

Figure 12.16

Restriction
enzymes
added
DNA sample 1

DNA sample 2

w
Cut
z
x

Cut

Cut

y

Longer
fragments

y

Sample
1

Sample
2
z

x
Shorter
fragments

w
y

y

12.17 Genomics is the scientific study of whole
genomes
 Genomics is the study of an organism’s complete
set of genes and their interactions.
– Initial studies focused on prokaryotic genomes.
– Many eukaryotic genomes have since been
investigated.


12.17 Genomics is the scientific study of whole
genomes
 Genomics allows another way to examine
evolutionary relationships.
– Genomic studies showed a 96% similarity in DNA
sequences between chimpanzees and humans.
– Functions of human disease-causing genes have been
determined by comparing human genes to similar
genes in yeast.


12.18 CONNECTION: The Human Genome
Project revealed that most of the human
genome does not consist of genes
 The goals of the Human Genome Project (HGP)
included
– determining the nucleotide sequence of all DNA in the
human genome and
– identifying the location and sequence of every human
gene.


12.18 CONNECTION: The Human Genome
Project revealed that most of the human
genome does not consist of genes
 Results of the Human Genome Project indicate that
– humans have about 20,000 genes in 3.2 billion
nucleotide pairs,
– only 1.5% of the DNA codes for proteins, tRNAs, or
rRNAs, and
– the remaining 98.5% of the DNA is noncoding DNA
including
– telomeres, stretches of noncoding DNA at the ends of
chromosomes, and
– transposable elements, DNA segments that can move or be
copied from one location to another within or between
chromosomes.

Figure 12.18

Exons (regions of genes coding for protein
or giving rise to rRNA or tRNA) (1.5%)

Repetitive
DNA that
includes
transposable
elements
and related
sequences
(44%)

Introns and
regulatory
sequences
(24%)

Unique
noncoding
DNA (15%)

Repetitive
DNA
unrelated to
transposable
elements
(15%)

12.20 Proteomics is the scientific study of the full
set of proteins encoded by a genome
 Proteomics
– is the study of the full protein sets encoded by
genomes and
– investigates protein functions and interactions.

 The human proteome includes about 100,000
proteins.
 Genomics and proteomics are helping biologists
study life from an increasingly holistic approach.


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