Download New New Developments Gene Therapy

New Developments
in
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New
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Developments Gene
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continuously.
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When can gene therapy be used?
Gene therapy is a difficult and complex process which is currently in
an early stage of development and is used in the treatment of a limited
number of medical conditions. Protein therapy has been successful for
many medical problems such as Pompe disease.
Originally, the particular therapeutic protein was extracted from
cadavers and abattoir material. Nowadays, with increased knowledge
and understanding of genetics and cell biology, it is possible to culture
cells that are genetically modified to produce a desired protein in
quantity.
Bacterial, plant, animal, yeast and human cells can be genetically
engineered by the insertion of a specific gene into their DNA. The gene
is then manipulated to remain active so the required protein is made
continuously. Chinese Hamster Ovary Cells are commonly used to
produce complex human proteins.
microinjection:
the DNA is injected by a tiny needle directly into the
host nucleus.
liposomes:
the DNA is carried in a ‘capsule of fat’ into the cell.
‘genetic shotgun’: microscopic metal particles coated with DNA are
shot into the cell.
electroporation:
a solution containing the target DNA and the host
cells is given a brief electric pulse that causes
temporary pores to open in the cell membrane and
increases the uptake of the target DNA.
Plasmid, virus and bacterial vectors are modified not to cause harm to
the host cell.
Bacterial host cells can take up plasmids from their culture solution.
in
in
in
New
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Developments
Developments Gene
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Therapy
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Fig. 3 Myozyme
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she was 6 weeks old and she is now on Myozyme.
Pompe disease manifests as a broad spectrum of clinical symptoms.
It affects fewer than 10,000 people worldwide. All patients typically
experience progressive muscle weakness and breathing difficulty,
but the rate of disease progression can vary widely depending on the
age of onset and the extent of organ involvement. When symptoms
appear within a few months of birth, babies frequently display a
markedly enlarged heart and die within the first year of life. When
symptoms appear during childhood, adolescence or adulthood,
patients may experience steadily progressive debilitation and
premature mortality due to respiratory failure. They often require
mechanical ventilation to assist with breathing and wheelchairs to
assist with mobility.
Myozyme is the first treatment ever approved for Pompe disease.
in
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New
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Our genes
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isolation,
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of around
33,00033,000
genes.genes.
Syllabus Reference
WhatWhat
makes
makes
us different?
us different?
Isolation
Isolation
Practical Activities
The DNA
The
containing
DNA containing
the normal
the human
normal gene
human
forgene
the specific
for the specific
protein is
protein is
Each person
Each
person
has almost
has 300
almost
different
300Biology
different
types oftypes
tissueof cell.
tissue
How
cell.
is How
it
is it removedremoved
from cultured
from cultured
human
human
cells and
cells
inserted
and inserted
into a DNA
vector:
a DNA vector: DNA from Kiwi Fruit
Leaving
Certificate
Practical
Activity
-intoExtracting
that these
thattissue
thesecells
tissue
arecells
different
are different
in structure
in structure
if they are
if they
genetically
are genetically a virus, bacterium
a virus, bacterium
or liposome
or(a
liposome
(fat
capsules).
(fat
capsules).
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vector
carry
can
it into
carry it into
home
experiment)
identical?
identical?
Each type
Each
of type
cell uses
of cell uses
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cellhost
where
cellitwhere
becomes
it becomes
incorporated
incorporated
into the into
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DNA.
host DNA.
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structure
DNA
structure
2.5.2subset
Gene Expression
a different
aUnit
different
subset
of– Heredity
our 33,000
of ourand
33,000
Cool 100 ml of methylated spirit in a freezer or using a salt-ice mixture.
In the case
In the
of case
bacterial
of bacterial
and plants
and cells,
plantsplasmids
cells, plasmids
can alsocan
be also
usedbe used
genes. genes.
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Brain use
cellsabout
use about
as
vectors.
as
vectors.
Plasmids
Plasmids
are
small
are
pieces
small of
pieces
DNA
independent
DNAinindependent
theofcell’s
of
the cell’s
Dissolve
25
g ofof
salt
900ofml
water.
Add 80 ml of liquid detergent. Peel
Unit 2.5.3
– Genetic
Code
11,000 genes—far
11,000
genes—far
more
than
more
the
than the
main chromosome(s).
main chromosome(s).
Bacterial
plasmids
plasmids
are
circular.
are
Plasmids
multiply
a Bacterial
kiwi
fruit.
Mash
thecircular.
peeledPlasmids
fruit inmultiply
a bowl. Add about 100 ml of the salt/
3000 used
3000
byused
the average
by the average
human human
within the
within
cell the
andcell
there
andmay
there
be may
different
be different
types oftypes
themofinthem
the same
in the same
Unit
2.5.5are
– Protein
Synthesis
cell. Some
cell.
genes
Some
genes
used
arebyused
all by all
detergent solution. Mix well. Decant the liquid into a glass, discarding the
cell.
cell.
o
our cellsour
for cells
the everyday
for the everyday
operation
operation
pulp. Warm the glass for 150 minutes in tepid
water
(40
C).
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add
Fig.
2 Genzyme
Fig.
2 Genzyme
employees
employees
working
working
in manufa
i
Unitmaintenance,
2.5.9
– Genetic
Engineering
and maintenance,
and
such
as such
aerobic
as
aerobic
Transcription
Transcription
Transformation?
Transformation?
the
cold
methylated
spirits
down
the
inside
of
the
glass.
DNA
separates out
respiration.
respiration.
solution,
asisfine
ata theofjunction
between
the pulp solution
Expression
During During
transformation
transformation
the of
host
thecell
host
DNA
cell
DNA
cut white
by
is cut
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by
action
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action
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gene carries
the information
the information
the
cold
methylated
spirits.
restriction
restriction
nuclease
nuclease
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the
so that
foreign
the
DNA
foreign
is inserted
DNA is inserted
in
the correct
in the correctExpression
for the for
formation
the formation
of a particular
of a particular
Expression
by the genetically-modified
by the genetically-modified
cells in
location.location.
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The are
pieces
joined
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joined
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the action
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ligase.
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so that
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are
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stages
two
in
stages
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transformation
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the
transformation
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and
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and
the
variety variety
of functions.
of functions.
Many of
Many of
Practical
Activity
Making
a
Model
of
DNA
Once
the
Once
genetically-modified
the
genetically-modified
tissue
culture
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is
c
uptake
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of
the
gene.
human
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gene.
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put in
beplace
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to e
t
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at specific
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restriction
sites— are
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are obtained
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that
that
catalysecatalyse
biochemical
biochemical
Use
matchsticks,
coloured
plasticine
and
other
suitable
materials
to
build
ProteinProtein
synthesis
synthesis where they
active form.
activeToform.
ensure
To ensure
that there
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no che
is
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form
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of the
partbacterial
of the bacterial
defencedefence
against against
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within the
within
cell.theSome
cell. Some
three models of each of the four DNA contamination,
nucleotides
each
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chromatography
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are structural
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to used
purify
purify
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the
protein.
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of prod
Transfection
Transfection
phosphate group, deoxyribose (a pentose used
sugar)
andtoathe
base.
Make
short
On completing
this
lesson the student should be
able
to:
membranes,
membranes,
tendons,tendons,
ligaments,
ligaments,
organisms
organisms
is referred
is referred
to as ‘pharming’.
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pairs simpler
of
complementary
hair, etc.
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Others
etc. Others
are involved
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in
Transfection
Transfection
is an alternative
is an length
alternative
andof somewhat
and of
somewhat
method
method
of
of nucleotides.
• On
completing
lesson the student should be able to:
the transport
the
transport
of
fats,ofvitamins,
fats,this
vitamins,
Once
protein
the protein
is produced,
is produced,
the nextthe
challe
nex
introducing
introducing
transgenic
transgenic
DNA into
DNA
a cell.
intoItadoes
cell. not
It does
involve
not cutting;
involve cutting;
the
theOnce the
metals and
metals
and hormones
in theall
blood
in
thesomatic
blood
pharmed
therapeutic
therapeutic
protein reaches
protein reaches
the site ti
foreign foreign
DNA binds
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binds temporarily
to the host
to the
DNA.
hostIf DNA.
the cell
If the
divides,
cell divides,
the
thepharmed
• hormones
Explain
why
our
cells are genetically identical.
Practical Activity - Make a Model of
or act as
or receptors,
act as receptors,
antivirals
antivirals
or
or
it isDNA
needed.
it isReplication
needed.
There are
There
several
are several
methodsmethod
by wh
foreign DNA
foreign
is not
DNA
replicated.
is not replicated.
• Define and outline the functions of a gene.
antibodies.
antibodies.
example,
example,
some therapeutic
some therapeutic
proteinsproteins
have s
Learning Objectives
Make 12 more nucleotides as before using a different colour for the
Fig. 1 DNA
Fig.transcription
1 DNA transcription
and
and
typestypes
attachedattached
to them.toBacteria
them. Bacteria
do not have
do not
thehac
• and
Describe
how
a foreign gene
can
be inserted into a hostVector
cell’s Vector
DNA.
phosphates. Open the DNA model into twothese
separate
strands.
Following
the so plan
Genes
Genes
health
and
health
protein synthesis
protein synthesis
carbohydrate
these carbohydrate
parts and
parts
so and
yeast,
yea
Thereexpression
are
There
various
are various
vectors vectors
that
canthat
carry
can
out
carry
this
out
function:
this
function:
base
pair
rule,
build
a new
strand along each
strand
using
• depends
Define
the
following:
isolation,
transfection,
haveold
to have
be
used.
to be
used.the 12 newly
Our health
Our
health depends
on the
body’s
on the ability
body’s
toability
maketransformation,
to
the
make
required
the required
proteins
proteins
made
nucleotides.
plasmid
plasmid
vector:
vector:
the
recombinant
the
recombinant
plasmid
plasmid
DNA
is
DNA
takenisinto
taken
theinto the
and
gene
cloning.
correctly,
correctly,
at the right
at the
time
right
andtime
in the
andright
in the
quantity.
right quantity.
Failure on
Failure
just one
on just one
host cell.
host cell.
of the many
of thethousands
many thousands
proteins
of proteins
can cause
can very
cause
serious
very serious
medicalmedical
• Explain
theoforigin
of restriction
enzymes
and
their use in genetic
problems.
problems.
A faultyAgene
faultyis gene
the usual
is thereason
usual reason
for the for
specific
the specific
protein protein virus vector:
virus
vector:
the
virus
the
carries
virus recombinant
carries
recombinant
DNA- or
DNA
RNA or
into
RNA
the into
engineering.
Practical
Activity
Formation
of the
Messenger RNA (mRNA)
failure. In
failure.
many
Incases
manythe
cases
onlythe
possible
only possible
remedy remedy
is geneistherapy,
gene therapy,
in
in
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General Learning Points
True or False
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• The human genome is made up of about 33,000 genes.
• Different types of tissue cells use different subsets of the genes.
• Gene expression controls which proteins are made by a cell, when
they are made and in what quantity.
• Faulty genes lead to incorrect formation of specific proteins.
• Inserting the normal gene into a suitable host cell can produce the
correct protein.
• Genetically modified host cells are the basis of ‘pharming’.
• Tissue culture is an essential part of ‘pharming’ of non-bacterial host
cells i.e. cells that have nuclei.
c) A gene carries the information for a particular protein.
T F
d) An enzyme is a biological catalyst.
T F
e) It is not possible to genetically modify human cells.
T F
f) Plasmid vectors cannot be used to carry foreign
genes into human cells.
T F
g) A restriction enzymes cuts DNA at a specific location.
T F
h) Human cells make restriction enzymes.
T F
i) Bacteria can absorb DNA from their external environment.
T F
j) An active gene is said to be in a state of ‘expression’.
T F
k) All types of human protein can be made by genetically
modified bacteria.
T F
Check your answers to these questions on www.sta.ie
in
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New
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WhatWhat
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or liposome
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Answer
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following questions in relation to thisTransfection
experiment.
membranes,
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organisms
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is referred
is referred
to as ‘pharming’.
to as ‘pharming’.
hair, etc.
hair,
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in
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metals and
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forit his
work are
on
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antibodies.
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some
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Biographical Notes
Fig. 1 DNA
Fig.transcription
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and
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Genes
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typestypesbacterial DNA. During his work at Stanford attached
attached
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them.
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thehac
University
School
of medicine
these carbohydrate
these carbohydrate
parts and
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atcan
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DNA
(iv) Washing-up liquid is normally used in this experiment.There are
have to have
be used.
be used.
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Ourdepends
health depends
on the body’s
on the ability
body’stoability
maketo
the
make
required
the required
proteinsproteins
technologies
andplasmid
in 1975
called
forthe
ainto
temporary
halt on recombinant DNA
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is
its
function?
vector: vector:
the recombinant
the recombinant
plasmid
DNA
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taken
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taken
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correctly,
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at the
right
at the
time
right
and
time
in the
andright
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quantity.
right quantity.
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just one
on just one plasmidplasmid
research
host cell.
host cell. until further safety issues were addressed which resulted in the
of the many
of thethousands
many thousands
of proteins
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can is
cause
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cause
serious
very
serious
medical
medical
(v)
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chloride
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problems.
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usual reason
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publishing
of the
National
ofinto
Health
virus vector:the virus
the
carries
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DNA or Institutes
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into
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the guidelines. In 1991 Berg was
(vi)
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acases
protease
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failure. In
failure.
many
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possible
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gene therapy,
in
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appointed
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which the
which
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gene
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correct
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or the missing
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(vii)
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enzyme used in this experiment bacterium
Committee.
bacterium
vector: vector:
the crown
thegall
crown
bacterium
gall bacterium
‘injects’ ‘injects’
the recombinant
the recombinant
protein is
protein
administered
is administered
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to the patient.
plasmidplasmid
DNA into
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plant
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plant cells.
(viii) The final separation of the DNA involves the use of alcohol
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can
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can gene
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Ian Wilmut, born 1944
host nucleus.
host nucleus.
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Gene therapy
is a difficult
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in
In 1996, Wilmut, a Scottish embryologist, produced a cloned sheep (Dolly)
an earlyan
stage
earlyofstage
development
of development
and is used
and is
in used
the treatment
in the treatment
of a limited
of a limited
liposomes:
liposomes: the DNAthe
is carried
DNA is in
carried
a ‘capsule
in a ‘capsule
of fat’ into
of fat’
the into
cell.the cell.
from
differentiated
adult
mammalian
cells. This was the first successful
number2004
number
of medical
of medical
conditions.
conditions.
Protein therapy
ProteinOrdinary
therapy
has been
has
successful
been
successful
for
for
Leaving
Certificate
Level
many medical
many medical
problems
problems
such assuch
Pompe
as Pompe
disease.disease.
‘genetic‘genetic
shotgun’:
shotgun’:
microscopic
microscopic
metal
particles
metal
particles
coated coated
withlived
DNA
with
areDNA
aresix years. In 1973, Wilmut also
cloning
of
a
mammal.‘Dolly’
for
over
(i) What is meant by the term DNA profiling?
shot into
shot
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cell.the
the cell.
created
first ever calf (Frosty) from frozen embryos.
Originally,
Originally,
the particular
the particular
therapeutic
therapeutic
protein protein
was extracted
was extracted
from from
(ii)
State
two
uses of
DNANowadays,
profiling.
cadavers
cadavers
and abattoir
and abattoir
material.material.
Nowadays,
with increased
with increased
knowledge
knowledge electroporation:
electroporation:
a solution
a solution
containing
containing
the target
theDNA
target
and
DNA
theand
hostthe host
and understanding
and understanding
of genetics
of genetics
and celland
biology,
cell biology,
it is possible
it is possible
to culture
to culture
cells is cells
givenis agiven
brief aelectric
brief electric
pulse that
pulse
causes
that causes
cells that
cells
arethat
genetically
are genetically
modified
modified
to produce
to produce
a desired
a desired
protein protein
in
in
Read
about
famous
scientists at www.sta.ie
temporary
temporary
pores more
topores
open to
in
open
the cell
in other
the
membrane
cell membrane
and
and
quantity.quantity. For further examples of past paper questions
increases
increases
the uptake
the of
uptake
the target
of theDNA.
target DNA.
check
www.sta.ie
Bacterial,
Bacterial,
plant, animal,
plant, animal,
yeast and
yeast
human
and human
cells
can
cells
be can
genetically
be genetically Plasmid,Plasmid,
virus and
virus
bacterial
and bacterial
vectors vectors
are modified
are modified
not to cause
not toharm
cause
toharm to
engineered
engineered
by the insertion
by the insertion
of a specific
of a specific
gene into
gene
theirinto
DNA.
their
The
DNA.
gene
The gene the hostthe
cell.
host cell.
Fig. 3 Myozyme
Fig. 3 Myozyme
is used for
is used
the treatment
for the treatment
of Pomp
is then manipulated
is then manipulated
to remain
to active
remainso
active
the required
so the required
protein protein
is madeis made
host cells
host
can
cells
takecan
up take
plasmids
up plasmids
from their
from
culture
their solution.
culture solution.
Megan Assink
Meganwas
Assink
diagnosed
was diagnosed
with Pompe
with
continuously.
continuously.
ChineseChinese
HamsterHamster
Ovary Cells
Ovaryare
Cells
commonly
are commonly
used toused to BacterialBacterial
she wasshe
6 weeks
was 6old
weeks
and she
old and
is now
sheon
is My
no
produceproduce
complexcomplex
human proteins.
human proteins.
Did You Know?
•
In 1865 Gregor Mendel, an Austrian monk, first showed the inheritance
of traits of pea plants obeyed simple mathematical rules. Some traits
are more complex and are controlled by several genes. Nevertheless,
Mendel’s work laid the foundations of the science of genetics.
•
From his analysis Mendel defined what he called an allele as
the fundamental unit of heredity—or what is now called a gene.
(Today the word allele means a specific version of a gene.)
•
Mendel’s work was not widely known and its significance was not
appreciated until the twentieth century when it was rediscovered.
•
Within organisms, genetic information is mostly carried in
chromosomes, where it is coded in the chemical structure of particular
DNA (deoxyribonucleic acid) molecules.
Revise the Terms
Can you recall the meaning of these terms? Reviewing the
terminology is a powerful aid for recall and retention.
Genes; gene, somatic, receptor, gene therapy; Pompe Disease;
cadaver; abattoir; genetically modified; Chinese Hamster Ovary
Cells; isolation; transformation; expression; restriction nuclease;
DNA ligase, gene cloning; vector DNA; plasmid; cutting; uptake;
plasmid vector; transgenic; virus vector; recombinant; bacterium
vector; microinjection; liposome; genetic shotgun; electroporation;
pharming, parenterals.
Check the Glossary of Terms for this lesson at www.sta.ie