Download One cell with the recombinant plasmid

Biotechnology
Dolly and
surrogate Mom
Embryonic stem cells and
gene therapy
Genetically modified rice.
Biotechnology
Biotechnology is the engineering of organisms for useful purposes.
Usually involves the creation of hybrid genes and their introduction into organisms
in which some or all of the gene is not normally present.
Fourteen month-old genetically engineered (“biotech”) salmon (left) and
standard salmon (right).
Biotechnology
We’ll examine:
Animal cloning
Gene cloning for
pharmaceutical production
Genetically modified
foods
DNA fingerprinting
The promise and perhaps
perils of embryonic stem cells
Animal Cloning
Dolly and her surrogate mother 1997
Why Clone Animals?
Five genetically identical cloned pigs.
To answer questions of basic biology
For pharmaceutical production.
For herd improvement.
To satisfy our desires (e.g. pet
cloning).
Is Animal Cloning Ethical?
The first cloned horse and her surrogate
mother/genetic twin.
As with many important questions, the answer is beyond the scope of
science.
Video
1.
2.
3.
4.
Researchers
isolated a somatic
cell from an adult
female sheep
Transferred the
nucleus from that
cell to an egg cell
from which the
nucleus had been
removed
The egg cell, with
its new nucleus,
was behaving just
like a freshly
fertilized zygote
It developed into
an embryo, which
was implanted into
a surrogate mother
and carried to
term.
The Biotechnology
of Reproductive
Cloning
Saved by Cloning?
Article
Some are firm believers while
many view these approaches to be
more of a stunt.
Carbon Copy– the First Cloned Pet
(Science (2002) 295:1443)
The Next Step?
Attempts at human cloning are viewed very unfavorably in the scientific community.
Recombinant DNA, Gene Cloning, and Pharmaceutical Production
DNA can be cut at specific sequences using restriction enzymes.
This creates DNA fragments useful for gene cloning.
Restriction Enzymes are Enzymes That Cut DNA Only at
Particular Sequences
Restriction
enzyme
animation
The enzyme EcoRI cutting DNA at its recognition sequence
Different restriction enzymes have different recognition sequences.
This makes it possible to create a wide variety of different gene fragments.
Palindrome: word that is the same forward and backwords
Examples:
racecar
dad
mom
5'- G A A T T C -3‘
3'- C T T A A G -5'
DNA Cut by a Restriction Enzyme Can be Joined Together in
New Ways by Ligase
These are recombinant DNA and they often are made of DNA from
different organisms.
Plasmids are Used to Replicate a Recombinant DNA
Plasmids are small circles of DNA found in bacteria.
Plasmids replicate independently of the bacterial chromosome.
Pieces of foreign DNA can be added within a plasmid to create a
recombinant plasmid.
Replication often produces 50-100 copies of a recombinant
plasmid in each cell.
Harnessing the Power of Recombinant DNA Technology –
Human Insulin Production by Bacteria
Human Insulin Production by Bacteria
and cut with a restriction enzyme
6) join the plasmid and human fragment
Human Insulin Production by Bacteria
Mix the recombinant plasmid with
bacteria.
Screening bacterial cells to learn which contain the human insulin gene is the
hard part.
Route to the Production by Bacteria of Human Insulin
One cell with the
recombinant plasmid
This is the step when gene cloning takes place.
A fermentor used to grow
recombinant bacteria.
The single recombinant plasmid replicates within a cell.
Then the single cell with many recombinant plasmids produces trillions of
like cells with recombinant plasmid – and the human insulin gene.
Route to the Production by Bacteria of Human Insulin
The final steps are to collect the bacteria, break open the cells, and purify the
insulin protein expressed from the recombinant human insulin gene.
Route to the
Production by
Bacteria of
Human Insulin
Overview of gene
cloning.
Cloning
animation
Transgenics
Transgenic organism: organism genetically
engineered by inserting a gene from another
organism
Transgenic animals, plants, bacteria
research, medical, and agricultural purposes
Transgenic Bacteria
Insulin, growth hormones, and substances that
dissolve blood clots
Transgenic bacteria slow formation of ice
crystals on crops to prevent from crop
damage, clean up oil spills, and decompose
garbage
Transgenic Plants
Genetically engineered to have more
resistance to insect or viral pests
Herbicide and insecticide resistant soybeans,
corn, cotton and canola
Peanuts and soybeans that do not cause
allergic reactions
Rice plants with increased iron and vitamins
to decrease malnutrition
Transgenic Animals
Transgenic livestock produced to improve
food supply and human health
Transgenic goats engineered to secrete a
protein called antithrombin III which
prevents human blood from forming clots
during surgery
Pharming
Pharming is the production of pharmaceuticals in animals engineered to
contain a foreign, drug-producing gene.
These goats contain the human gene for a clot-dissolving
protein that is produced in their milk.
Protein
Animal
Use
Antithrombin III
Goat
Reduce amount of blood
needed in some surgeries
Factor VIII, Factor IX
Goat, Pig, Sheep
Treatment of Hemophilia
CFTR
Sheep
Treatment of cystic
fibrosis
Lactoferrin
Cow
Natural antibiotic and
used in coronary surgery
Alpha-1-antitrypsin
Sheep
Treatment of cystic
fibrosis and emphysema
Lysostaphin
Cow
An anti-bacteria
compound that prevents
mastitis in cows
Spider Silk Protein
Goat
Production of ultrastrong, lightweight
medical industiral
materials
The Guiness Book of Animal Records 1995.
"Spider silk is the strongest of all natural and
man-made fibres.... It is even stronger than
steel: the dragline of a European garden spider
(Araneus diadematus), for example, can
support a weight of 0.5 g (0.002 oz) without
snapping, whereas a steel strand of similar
thickness will snap under the strain of just
0.25 g (0.01 oz)."
"Spider silk is extremely strong -- it is about five times stronger than steel and
twice as strong as Kevlar of the same weight. Spider silk also has the ability to
stretch about 30-percent longer than its original length without breaking, which
makes it very resilient."
The Stem Cell Concept
A stem cell is an undifferentiated,
dividing cell that gives rise to a
daughter cell like itself and a
daughter cell that becomes a
specialized cell type.
The Promise and Possible Perils of Stem Cells
Stem Cells are Found in the Adult, but the Most Promising Types of
Stem Cells for Therapy are Embryonic Stem Cells
The Inner Cell Mass is the Source of Embryonic Stem Cells
The embryo is destroyed by separating it into individual cells for the collection
of ICM cells.
Some Thorny Ethical
Questions
Are these masses of cells a
human?
Is it ethical to harvest
embryonic stem cells from
the “extra” embryos created
during in vitro fertilization?
DNA, the Law, and Many Other Applications –
The Technology of DNA Fingerprinting
A DNA fingerprint used in a murder case.
The defendant stated that the blood on his clothing was his.
What are we looking at? How was it produced?
DNA Fingerprinting Basics
Different individuals carry different alleles.
Most alleles useful for DNA fingerprinting differ on the basis of the
number of repetitive DNA sequences they contain.
DNA Fingerprinting Basics
If DNA is cut with a restriction
enzyme that recognizes sites on either
side of the region that varies, DNA
fragments of different sizes will be
produced.
DNA samples treated with ethidium
bromide are inserted into wells in a
piece of agar (gel)
When a charge is applied, DNA
which is negative will travel to the
positive end of the gel
Smaller pieces will travel the
furthest
Bands are visualized under UV light
A Site With Three Alleles Useful for DNA Fingerprinting
DNA fragments of different size will be produced by a restriction
enzyme that cuts at the points shown by the arrows.
The DNA Fragments Are Separated on the Basis of Size
The technique is called gel electrophoresis.
The pattern of DNA bands is compared between each sample loaded on
the gel.
Gel electrophoresis animation
Possible Patterns for a Single “Gene” With Three Alleles
In a standard
DNA
fingerprint,
about a dozen
sites are
analyzed, with
each site
having many
possible
alleles.
A DNA Fingerprint
When many genes are
analyzed, each with many
different alleles, the
chance that two patterns
match by coincidence is
vanishingly small.
DNA detective
animation
HGP fingerprinting
page
DNA and the Law
SLT 3/8/05
Some applications of DNA
fingerprinting in the justice system.
Restriction Enzymes and Gel
Electrophoresis
http://arbl.cvmbs.colostate.edu/hbooks/genetic
s/biotech/gels/virgel.html
Biotechnology Australia site - Games
Genetically Modified Foods
Many of our crops in the US are genetically modified.
Should they be?
GM Crops are Here Today
Source: Pew Initiative on Food and Biotechnology, August 2004.
Methods for Plant Genetic Engineering are Well-Developed and
Similar to Those for Animals
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2.
3.
Restriction enzyme cut plasmid
Insert gene of interest
Introduce to plant cells
Golden Rice is Modified to be Provide a Dietary Source of Vitamin A
Golden rice (yellow)
with standard rice
(white).
Worldwide, 7% of children suffer vitamin A deficiency, many of them living in
regions in which rice is a staple of the diet.
Genetically Modified Crops
Genetically Modified Cotton
(contains a bacterial gene for
pest resistance)
Standard Cotton
GMOs, Especially Outside the US, Are a Divisive Issue
Protesters at the 2000 Montreal World
Trade Summit
European sentiment
Current Concerns by Scientists Focus on Environmental, Not
Health, Effects of GM Crops
The jury’s still out on the magnitude of GM crop’s ecological impact, but the
question is debated seriously.
Current Concerns
by Scientists
Focus on
Environmental,
Not Health,
Effects of GM
Crops