Ch. 13 Genetic Engineering

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14 Δεκ 2012 (πριν από 4 χρόνια και 7 μήνες)

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Biotechnology and

Genetic Engineering

Outline


Biotechnology: An introduction



Changing the Living World


Selective Breeding


Increasing Variation



Manipulating DNA


The Tools of Molecular Biology


Using the DNA Sequence

Outline


Cell Transformation


Transforming Bacteria


Transforming Plant Cells


Transforming Animal Cells



Applications of Genetic Engineering


Transgenic Organisms


Cloning

Introduction


Biotechnology

is essentially


the use of
living

organisms (often
minute

microorganisms) and
their
products



for health, social or economic
purposes.


Biotechnology is widely considered to be
the
growth

technology of the
21st
Century

which will lead to huge growth
in the Biotechnology industry and exciting
opportunities

for graduates

Introduction


Through genetic engineering scientists
can combine DNA from different sources
and this process is called
“R
ecombinant DNA technology”


The secrets of DNA structure and
functions have led to
gene cloning

and
genetic engineering
, manipulating the
DNA of an organism


Introduction




Biotechnology

is an
interdisciplinary

science including not only
biology
, but also
subjects like
mathematics, physics, chemistry
and engineering
.



It is a blend of various technologies applied together
to living cells for
production

of a particular
product

or for
improving

upon it.

Application of Biotechnology


Its use and application ranges from fields
like
agriculture to industry

(food,
pharmaceutical, chemical,
bioproducts
,
textiles etc.),
medicine, nutrition,
environmental conservation, animal
sciences

etc. making it one of the fastest
growing fields.


The work is generally carried out in the
laboratories, as it is a scientific research
oriented field.

Application of Biotechnology

Applications of biotechnology are widespread,
including the following:


diagnosis and treatment of human diseases.


improved production of therapeutic agents.


development of improved crop plant species.


development of improved pest/pathogen
control processes

Application of Biotechnology


development of biosensors for
environmental pollutants.


development of improved waste
treatment processes and methods for
remediation contaminated sites.


production of transgenic organisms for
production of new drugs, improved
transplantation success and improved
animal and plant.






Biotechnology's application in this area
has helped in the development of
various medicines, vaccines for
various diseases and diagnostics.



These include


Transplant Organs for Humans


Tissue Engineering


Biopharmaceuticals



MEDICINE AND HEALTH CARE

AGRICULTURE AND ANIMAL
HUSBANDRY


Genetic engineering has led to genetically
modified crops and animals that are more
hardy and resistant to extreme conditions



These genetically modified organisms
(
GMOs
) also have other improvements
such as faster growth, bigger yield, and
higher nutrient content.



Through biotechnology, researchers are
making meat and dairy products better
to eat.


They are improving milk so that 90
percent of adults who are lactose
-
intolerant can drink it.


They also are identifying genes that will
result in less fat in beef and improved
flavor and tenderness.




Scientists are trying to
find genes that cause or
resist certain diseases of
livestock, such as "mad
cow disease“.



Once these genes are
found in livestock,
scientists can breed
livestock for the resistant
gene and develop
vaccines and diagnostic
tests to prevent these
diseases
.

Selective Breeding


Humans use selective
breeding, which takes
advantage of naturally
occurring genetic
variation in plants,
animals, and other
organisms, to pass
desired traits on to the
next generation of
organisms

Selective Breeding


Breed only those
plants or animals
with desirable traits



People have been
using selective
breeding for 1000’s
of years with farm
crops and
domesticated
animals.


Selective Breeding


Nearly all
domestic
animals

--

including
horses,
cats, and
farm animals



and
most
crop plants

have been produced
by selective breeding

No freaking way!

Hybridization


Louis Burbank was the
greatest selective
breeder of all time. He
developed the
disease
-
resistant
potato

and more than
800
varieties of
plants.


Louis Burbank used the technique
of
hybridization

and bred
dissimilar individuals
to combine
the best traits of both parents
.



The
hybrids

produced by these
crosses were
hardier

than their
parents

Inbreeding


To
maintain the desired
characteristics

of a line of
organisms, breeders often
use the technique of
inbreeding.



Inbreeding

is the
continued breeding

of
individuals

with
similar
characteristics


Increasing Variation


In order for selective breeding to be
successful, there must be a lot of
genetic variation in the population



Breeders
increase

the genetic variation
in a population by
inducing
mutations
, which are the ultimate
source of genetic variability


Increasing Variation


Breeders increase the mutation rate by
using radiation and chemicals



The Basic Steps of Genetic Engineering

1.
Cutting the DNA
:


Restriction Enzymes
: bacterial enzymes that
recognize and bind
to specific short
sequences of DNA
, and then cut the DNA
between specific nucleotides within the
sequences.



Vector
:

agent used to
carry the gene
of
interest


usually plasmids



Plasmid
: the circular DNA molecules that
replicate

The Basic Steps to Genetic Engineering

2.
Making Recombinant DNA


DNA fragments of interes
t (that have
already been cut) are combined with the
vector.


DNA
ligase



the enzyme bonds the 2 ends of
the fragments to the vectors.


3.
Cloning


Gene cloning
: the process of making many
copies of a gene


Bacteria reproduce by binary fission

The Basic Steps to Genetic
Engineering

4.
Screening


Cells that
have received the gene
of
interest
are separated out
.


Those cells then continue to produce the
protein coded for by the gene

Cutting DNA & Making Recombinant
DNA


How Restriction enzymes work:


The Enzymes
recognize

specific
sequences

on
Human and Bacterial Plasmids


The Enzymes
cut the strands
.


The
cut

produces DNA
fragments

with
short
strands on each end

that are complementary to
each other


“Sticky Ends”


Both the human
DNA and the Plasmid “Open Up

with the same sticky ends remaining


They Bind Together


Diagram

Recognition
sequences

DNA sequence

Recognition sequences

DNA sequence

Restriction enzyme

EcoR
I
cuts
the DNA into fragments.

Sticky end

Confirmation of a Cloned Gene


One method used identify a specific gene
is called a
Southern Blot


Steps:

1.
Cut
DNA from bacteria

with restriction
enzymes.

2.
DNA fragments
are separated by a gel
soaked in a chemical solution.


Gel electrophoresis


uses an electric field
within a gel to separate molecules by their size

Confirmation of a Cloned Gene


Negatively charged DNA
is put into
these wells.


They are
attracted to the positive

pole
from the electric field.



The
Smallest

DNA fragments move the
fastest


Gel Electrophoresis

DNA plus
restriction enzyme

Mixture of
DNA
fragments

Gel

Power source

Longer
fragments

Shorter
fragments

Confirmation of a Cloned Gene

3.
The
DNA

separated is then transferred to a
filter paper (blotted) and a probe solution is
added.


Probes:

radioactive RNA

or single
-
stranded
DNA pieces that are complementary to the gene
of interest


4.
Only DNA fragments complementary to the
probe will form and bind bands

Recombinant Bacteria

1.
Remove bacterial DNA
(plasmid).


2.
Cut the Bacterial DNA with

restriction enzymes
”.


3.
Cut the DNA from another
organism with “
restriction
enzymes”.


4.
Combine the cut pieces of DNA
together with another enzyme
and insert them into bacteria.


5.
Reproduce the recombinant
bacteria.


6.
The foreign genes will be
expressed in the bacteria.



Confirmation of Cloned Genes



When a
bacteria or other cell

takes in a foreign piece of DNA such
as a plasmid, the process is called
transformation



If

transformation
is

successful,

the
recombinant
DNA is integrated

into one of
the chromosomes

of
the cell.



Creating HGH

Human Cell

Gene for human
growth hormone

Recombinant
DNA

Gene for human
growth hormone

Sticky
ends

DNA
recombination

DNA
insertion

Bacterial Cell

Plasmid

Bacterial
chromosome

Bacterial cell for containing gene
for human growth hormone

Some Benefits
of Recombinant
Bacteria

1.
Bacteria can make human insulin or
human growth hormone.


1.
Bacteria can be engineered to “eat” oil
spills.

The DNA of plants and animals


can also be altered.


PLANTS


1.
disease
-
resistant and
insect
-
resistant crops


2. Hardier fruit


3. 70
-
75% of food in
supermarket is
genetically modified
.



Improving Crops


Genetic engineers can
add favorable
characteristics

to a plant



Plants become resistant to
insects

(no longer need pesticides); resistant
to
weed killer

(so crops won’t die,
but weeds will); improved nutrition


rice + corn

Plant Transformation

Recombinant
plasmid

Gene to be
transferred

Agrobacterium
tumefaciens

Cellular
DNA

Transformed bacteria introduce
plasmids into plant cells

Plant cell
colonies

Complete plant is
generated from
transformed cell

Inside plant cell,
Agrobacterium
inserts part of its
DNA into host cell
chromosome

How to Create a Genetically

Modified Plant

1.Create
recombinant
bacteria with
desired gene.


2. Allow the
bacteria to
“infect" the
plant cells.


3. Desired gene is
inserted into
plant
chromosomes.

Genetically modified organisms
are called
transgenic organisms
.


TRANSGENIC ANIMALS


1.
Mice


used to study human
immune system


2.
Chickens


more resistant to
infections


3.
Cows


increase milk supply
and leaner meat



4. Goats, sheep and pigs


produce human proteins in
their milk




Human DNA in
a Goat Cell

This goat contains a human
gene that codes for a blood
clotting agent. The blood
clotting agent can be harvested
in the goat’s milk.

.

Transgenic Goat

Transgenic Cows


Growth hormones

are given to cows to
produce more milk



Human genes are added to farm animals in
order to have
human proteins in their
milk


The Human
proteins

are
extracted

from
milk and
sold to pharmacy companies
.



Useful for complex proteins that can’t
be made in bacteria

Transgenic animals


Animals that have foreign DNA in
their cells



Cloning of animals is another way
to make large quantities of a
certain protein.


Transgenic animals



How it works: an
intact nucleus

from an
embryonic cell (whose DNA has recombined
with a human gene) is
placed

into an
egg

whose nucleus has been removed.



The “
new


egg

is then placed into the
uterus of an animal.

How to Create a

Transgenic Animal

Animal Cloning


A
clone

is a member of a population of
genetically identical cells

produced from
a single cell



How it works: an
intact nucleus

from a cell
is removed

Cloning Animals


The nucleus is
fused

with a egg cell (whose
nucleus has been removed) taken from
another adult



The fused cell begins to divide and the
embryo is placed in the uterus
of a
foster mother.



The “
new


egg

is then develops normally.

Cloning

A donor cell is taken from a
sheep’s udder
.

Donor Nucleus

These two cells are fused using an
electric shock.

Fused Cell

The fused cell begins dividing
normally.

Embryo

The embryo is placed in the uterus of
a foster mother.

Foster Mother

The embryo develops normally into a lamb

Dolly

Cloned Lamb

Egg Cell

An egg cell is taken from an
adult female sheep.

The nucleus of the egg cell is removed.


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Genetically Engineered Drugs and
Vaccines


Today, many pharmaceutical companies
around the world produce
important
proteins
using genetic engineering.



Vaccine
: a solution containing all or part of
a
harmless version of a pathogen
; used
to prevent viral diseases (don’t respond to
drugs)



Many vaccines are made using genetic
engineering