Download DNA Technology ppt chapter 13 Honors Txtbk

Teresa Audesirk • Gerald Audesirk • Bruce E. Byers
Biology: Life on Earth
Eighth Edition
Lecture for Chapter 13
Biotechnology
Copyright © 2008 Pearson Prentice Hall, Inc.
Chapter 13 Outline
•
•
•
•
•
13.1 The World of Biotechnology, p. 252
13.2 DNA Recombination in Nature, p. 252
13.3 Biotechnology in Forensics, p. 254
13.4 Biotechnology in Agriculture, p. 258
13.5 Biotechnology and the Human Genome, p.
261
• 13.6 Biotechnology in Medicine, p. 262
• 13.7 Biotechnology and Ethics, p. 265
Section 13.1 Outline
• 13.1 The World of Biotechnology
– Traditional Applications of Biotechnology
– Genetic Engineering
– Recombinant DNA
Traditional Applications
•
Biotechnology is applied biology
– Modern focus on genetic engineering,
recombinant DNA technology, and analysis of
biomolecules
Traditional Applications
•
Traditional (historical) applications of
biotechnology date back to over 10,000
years ago
– Use of yeast to produce beer and wine in Egypt
and Near East
– Selective breeding of plants
– Selective breeding of animals
Genetic Engineering
•
Genetic engineering refers to the
modification of genetic material to achieve
specific goals
Genetic Engineering
•
Major goals of genetic engineering
– Learn more about cellular processes,
including inheritance and gene expression
– Provide better understanding and treatment of
diseases, particularly genetic disorders
– Generate economic and social benefits
through production of valuable biomolecules
and improved plants and animals for
agriculture
Recombinant DNA
•
Genetic engineering utilizes recombinant
DNA technology
– Splicing together of genes or portions of
genes from different organisms
Recombinant DNA
•
Recombinant DNA can be transferred to
plants and animals
– Modified animals are called transgenic or
genetically modified organisms (GMOs)
– Most modern biotechnology includes
manipulation of DNA
Section 13.2 Outline
•
13.2 DNA Recombination in Nature
– Sexual Reproduction Recombines DNA
– Transformation: Acquisition of DNA from the
Environment
– Viral Transfer: Movement of DNA Between
Organism via Viruses
Recombination in Nature
•
Many natural processes recombine DNA
Sexual Reproduction
•
Due to crossing over during meiosis, each
chromosome in a gamete contains a
mixture of alleles from the two parental
chromosomes
– Thus, eggs and sperm contain recombinant
DNA
Transformation
•
Bacteria can naturally take up DNA from
the environment (transformation) and
integrate the new genes into the genome
(recombination)
Transformation
•
Small circular DNA molecules (plasmids)
carry supplementary genes
– Plasmid genes may allow bacteria to grow in
novel environments
– Plasmid genes may enhance virulence of
bacteria in establishing an infection
– Plasmid genes may confer resistance to
antimicrobial drugs
Viral Transfer of DNA
•
Viral life cycle
1.
2.
3.
4.
Viral particle invades host cell
Viral DNA is replicated
Viral protein molecules are synthesized
Offspring viruses are assembled and break
out of the host cell
Viral Transfer of DNA
•
Viral transfer of DNA
– Viruses may package some genes from host
cell into viral particles during assembly
– Infection of new host cell injects genes from
previous host, allowing for recombination
Section 13.3 Outline
•
13.3 Biotechnology in Forensics
– How Biotechnology Revolutionized Forensics
– Amplification of DNA by Polymerase Chain
Reaction
– Gel Electrophoresis: Separation of DNA
Fragments
– DNA Probes Are Used to Highlight Bands in a
Gel
– DNA Fingerprinting
Biotechnology and Forensics
• Forensics is the science of criminal and
victim identification
Biotechnology and Forensics
• DNA technology has allowed forensic
science to identify victims and criminals from
trace biological samples
– Genetic sequences of any human individual are
unique
– DNA analysis reveals patterns that identify
people with a high degree of accuracy
Polymerase Chain Reaction
•
•
Forensic technicians typically have very
little DNA with which to perform analyses
Polymerase Chain Reaction (PCR)
produces virtually unlimited copies of a
very small DNA sample
Polymerase Chain Reaction
•
•
PCR requires small pieces of DNA (called
primers) that are complementary to the
gene sequences targeted for copying
A PCR “run” is basically DNA replication
in a tiny test tube
– Template DNA, primer, nucleotides, and DNA
polymerase are all in the reaction mix
Polymerase Chain Reaction
•
Four steps of a PCR cycle
1. Template strand separation
– The test tube is heated to 90-95oC to cause the
double stranded template DNA to separate into
single strands…
Polymerase Chain Reaction
•
Four steps of a PCR cycle
2. Binding of the primers
– The temperature is lowered to 50oC to allow the
primer DNA segments to bind to the targeted gene
sequences through hydrogen bonding…
Polymerase Chain Reaction
•
Four steps of a PCR cycle
3. New DNA synthesis at targeted sequences
– The temperature is raised to 70-72oC where the
heat-stable DNA polymerase synthesizes new
DNA of the sequences targeted by the primers…
Polymerase Chain Reaction
•
Four steps of a PCR cycle
4. Repetition of the cycle
– The cycle is repeated automatically (by a
thermocycler machine) for 20-30 cycles,
producing up to 1 billion copies of the original
targeted DNA sequence
Polymerase Chain Reaction
•
•
Choice of primers determines which
sequences are amplified (copied)
Forensic scientists focus on short
tandem repeats (STRs) found within the
human genome
Polymerase Chain Reaction
•
•
•
STRs are repeated sequences of DNA
within the chromosomes that do not code
for proteins
STRs vary greatly between different
human individuals
A match of 10 different STRs between
suspect and crime scene DNA virtually
proves the suspect was at the crime
scene
Gel Electrophoresis
•
•
Mixtures of DNA fragments can be
separated on the basis of size
Gel electrophoresis is a technique used
to spread out different-length DNA
fragments in a mixture
Gel Electrophoresis
•
Four steps of gel electrophoresis
1. DNA mixtures are placed into wells at one
end of a slab of agarose gel
Gel Electrophoresis
•
Four steps of gel electrophoresis
2. An electric current introduced through the gel
causes the negatively-charged DNA
fragments to migrate towards the positive
electrode
Gel Electrophoresis
•
Four steps of gel electrophoresis
3. Short DNA fragments move more easily
through the three-dimensional meshwork of
fibers between the gel
– Short DNA fragments migrate farther than long
DNA fragments so the mixture is separated into
bands of DNA of specific lengths
Gel Electrophoresis
•
Four steps of gel electrophoresis
4. The invisible bands of DNA are made visible
using stains or DNA probes
DNA Probes
•
DNA probes are short single-stranded
DNA fragments used to identify DNA in a
gel pattern
– Probe sequence is complementary to a DNA
fragment somewhere in the gel pattern
DNA Probes
•
DNA probes
– Probes identify the location of a gene
sequence by hydrogen-bonding to the band
containing it
DNA Probes
•
DNA probes
– Probes may have colored molecules
attached to them to allow for visual
identification of the bands to which they bind
– Gel DNA pattern is usually transferred to
piece of nylon paper before probing
DNA Fingerprinting
•
•
DNA from a crime scene sample can be
amplified by PCR and run on a gel with
suspect DNAs
Short tandem repeats (STRs) in the gel
DNA can be identified by DNA probes
DNA Fingerprinting
•
•
Distinctive pattern of STR numbers and
lengths are fairly unique to a specific
individual (forming a DNA fingerprint)
DNA fingerprint from crime scene can be
matched with DNA fingerprint of suspect
Section 13.4 Outline
• 13.4 Biotechnology in Agriculture
– Many Crops Are Genetically Modified
– Cloning of the Desired Gene
– Restriction Enzymes Cut DNA at Specific Places
– Splicing of DNA Fragments Is Aided by Sticky
Ends
Section 13.4 Outline
• 13.4 Biotechnology in Agriculture
(continued)
– Plasmids Are Used to Insert Genes Into a Plant
Cell
– GM Plants May Be Engineered to Produce
Medicines
– GM Animals May Be Useful in Agriculture and
Medicine
Many Crops Are Genetically Modified
•
One third to three-quarters of corn, cotton,
and soybeans grown in the US are
genetically modified
Many Crops Are Genetically Modified
•
Crop plants are commonly modified to
improve insect and herbicide resistance
– Herbicide resistant crops withstand
applications of weed-killing chemicals
– Bt gene (from Bacillus thuringiensis
bacterium) can be inserted into plants to
produce insect-killing protein in crops
Cloning of the Desired Gene
•
Modifying a plant genetically begins with
gene cloning
1. Desired gene is first isolated from organism
containing it
•
Desired gene may alternately be synthesized in
the laboratory
Cloning of the Desired Gene
•
Modifying a plant genetically begins with
gene cloning
2. Gene is next inserted into a small DNA circle
called a plasmid which replicates itself
autonomously in bacterial cells
Restriction Enzymes Cut DNA
•
•
A DNA sequence (e.g. a gene) can be
removed from a chromosome using special
enzymes
Restriction enzymes are nucleases that
cut DNA at specific nucleotide sequences
Restriction Enzymes Cut DNA
•
Enzymes that create staggered cuts with
“sticky ends” are the most useful in gene
cloning
Splicing of DNA Fragments
•
Sticky ends allow for splicing of a DNA
fragment with another complementary
fragment
– Bt gene can be cut out of the Bacillus
chromosome with the same enzyme used to
cut open the plasmid
– Bt gene fragment ends can base-pair with
sticky ends of the opened plasmid, adding
gene to the plasmid circle
Splicing of DNA Fragments
•
DNA ligase enzyme used next to
permanently bond gene into plasmid
Plasmids Are Used to Insert Genes
•
The Ti plasmid from Agrobacterium
tumefaciens is ideal for transferring genes
into plant chromosomes
Plasmids Are Used to Insert Genes
•
Agrobacterium infects plant cells and
inserts its small Ti plasmid into a plant
chromosome in the nucleus
– Pathogenic effects of certain tumor-causing Ti
plasmid genes can be disabled
– A gene inserted into a Ti plasmid is therefore
carried into the plant cell chromosomes by a
natural process
GM Plants and Medicines
•
Medically useful genes can be inserted
into plants—example:
– Potatoes have been engineered to produce
harmless hepatitis B virus and E. coli proteins,
stimulating an immune response when eaten
GM Plants and Medicines
•
Medically useful genes can be inserted
into plants—example:
– Plants could be engineered to produce human
antibodies, conferring passive immunity to
microbial infection merely by eating the plant
GM Animals
•
Transgenic (genetically modified or GM)
animals can be engineered by incorporating
genes into chromosomes of a fertilized egg
GM Animals
•
Healthy transgenic animals are difficult to
engineer
– Growth hormone genes have been inserted
into pigs and fish species but some
abnormalities have been observed
•
Animals like sheep might be engineered to
produce medically important proteins in
their milk
Section 13.5 Outline
•
13.6 Biotechnology and the Human
Genome
– Findings and Applications of the Human Genome
Project
The Human Genome Project
•
Findings
– Human genome contains ~25,000 genes
– New genes, including many disease-associated
genes have been discovered
– Has determined the nucleotide sequence of all
the DNA in our entire set of genes, called the
human genome
– The genes comprise 2% of all the DNA
The Human Genome Project
•
Applications
– Improved diagnosis, treatment and cures of
genetic disorders or predispositions
– Comparison of our genome to those of other
species will clarify the genetic differences that
help to make us human
Section 13.6 Outline
•
13.6 Biotechnology in Medicine
– DNA Technology Can Be Used to Diagnose
Inherited Disorders
– Restriction Enzyme Fragment Analysis
– Identification of Defective Alleles with DNA
Probes
– DNA Technology Can Be Used to Treat
Disease
Diagnosis of Inherited Disorders
•
•
Potential parents can learn if they are
carriers of a heritable disorder through
testing
Alleles for defective genes differ from
normal, functional genes in nucleotide
sequence
Diagnosis of Inherited Disorders
•
Two methods employed to detect a
defective allele in a person’s DNA sample
– Restriction enzyme fragment analysis
– Identification of defective alleles with DNA
probes
Restriction Enzyme Fragment Analysis
•
A particular restriction enzyme may cut
two different alleles of a gene differently
– Differences in nucleotide sequence within
genes produces different numbers of cutting
sites and different lengths of fragments
Restriction Enzyme Fragment Analysis
– Differences in restriction enzyme fragments
between genes are known as restriction
fragment length polymorphisms (RFLPs)
– RFLP differences are revealed in gel
electrophoresis
Restriction Enzyme Fragment Analysis
•
•
Before PCR, restriction fragment
differences were used to generate DNA
fingerprints in forensics
RFLP analysis is now commonly used to
identify the presence of the sickle-cell
anemia gene in a person’s DNA
DNA Probes
•
•
Defective alleles can also be identified
using DNA probes
DNA probing is especially useful where
there are many different alleles at a single
gene locus
– Cystic fibrosis is a disease caused by any of
32 alleles out of 1000 total possible alleles
DNA Probes
•
Arrays of single-stranded DNA
complementary to each of the defective
alleles can be bound to filter paper
1. A person’s DNA sample is cut up and separated
into single-strands
2. The array is bathed in the DNA sample
3. Strands of DNA binding to complementary
sequence on the paper indicate presence of a
defective allele in person’s genome
DNA Probes
•
•
•
An expanded version of this type of DNA analysis
is known as a microarray
A microarray contains up to thousands of probes
for a variety of disease-related alleles
Microarray analysis has the potential to
comprehensively identify disease susceptibility
Disease Treatment
•
Treatments using DNA technology
– Administration of proteins to treat but not cure
a disorder
• Human insulin produced inexpensively and
rapidly in recombinant bacteria for
diabetics
• Growth hormone and blood clotting factors
produced safely and inexpensively in
recombinant bacteria
Disease Treatment
•
Treatments using DNA technology
– Replacing defective genes to possibly cure a
disorder
• Replacement of defective cystic fibrosis
allele using a virus to carry in a functional
gene sequence into patient lung cells
• Defective bone marrow cell DNA
replacement by functional gene in severe
combined immune deficiency (SCID)
patients
Section 13.7 Outline
•
13.7 Biotechnology and Ethics
– Issues Surrounding GM Organisms in
Agriculture
– Scientific Objections to Genetically Modified
Organisms
– Ethics of Using Biotechnology on the Human
Genome
GM Organisms in Agriculture
•
The goal of breeding or genetically
modifying plants or livestock is to make
them more productive, efficient, or useful
GM Organisms in Agriculture
•
Genetic modification differs from selective
breeding (“traditional biotechnology”)
– Genetic engineering is much more rapid
– Genetic engineering can transfer genes
between species
– Genetic engineering can produce new genes
never seen before on Earth
GM Organisms in Agriculture
•
Benefits of genetically modified plants
– Transgenic crops decrease applications of
pesticides, saving fuel, labor, and money
– GM plants can be sold at a lower price due to
farm savings
– Genetically engineered crops can deliver
greater amounts of vitamins
•
e.g. “golden rice” which produces vitamin A
Scientific Objections to GMOs
•
Safety issues from eating GMOs
– Could ingestion of Bt protein in insect-resistant
plants be dangerous to humans?
– Are transgenic fish producing extra growth
hormone dangerous to eat?
Scientific Objections to GMOs
•
Safety issues from eating GMOs
– Could GM crops cause allergic reactions?
•
USDA now monitors GM foods for allergic potential
– Toxicology study of GM plants (2003)
concluded that ingestion of current transgenic
crops pose no significant health dangers
Scientific Objections to GMOs
•
Environmental hazards posed by GMOs
– Pollen from modified plants can carry GM
genes to the wild plant population
•
Could herbicide resistance genes be transferred to
weed species, creating superweeds?
Scientific Objections to GMOs
•
Environmental hazards posed by GMOs
– Could GM fish reduce biodiversity in the wild
population if they escape?
•
Reduced diversity in wild fish makes them more
susceptible to catastrophic disease outbreaks
Scientific Objections to GMOs
•
Environmental hazards posed by GMOs
– US found to lack adequate system to monitor
changes in ecosystem wrought by GMOs
(National Academy of Science Study 2003)
The Human Genome
•
Should parents be given information about
the genetic health of an unborn fetus?
The Human Genome
•
Should parents be allowed to select the
genomes of their offspring?
– Embryos from in vitro fertilization are currently
tested before implantation
– Many unused embryos are discarded
The Human Genome
•
Should parents be allowed to design or
correct the genomes of their offspring?