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4.4.1 Outline the use of polymerase chain reactions (PCR) to copy and amplify
minute quantities of DNA
4.4.2 State that, in gel electrophoresis, fragments of DNA move in an electric field
and are separated according to their size
4.4.3 State that gel electrophoresis of DNA is used in DNA profiling
4.4.4 Describe the application of DNA profiling to determine paternity and also in
forensic investigations
4.4.5 Analyze DNA profiles to draw conclusions about paternity or forensic
investigations
4.4.6 Outline three outcomes of the sequencing of the complete human genome
4.4.7 State that, when genes are transferred between species, the amino acid
sequence of polypeptides translated from them is unchanged because the genetic
code is universal
4.4.8 Outline a basic technique used for gene transfer involving plasmids, a hose
cell (bacterium, yeast, or other cell), restriction enzymes (endonucleases) and DNA
ligase
4.4.9 State two examples of the current uses of genetically modified crops or
animals
4.4.10 Discuss the potential benefits and possible harmful effects of one example
of genetic modification
4.4.11 Define clone
4.4.12 Outline a technique for cloning using differentiated animal cells
4.4.13 Discuss the ethical issues of therapeutic cloning in humans
 DNA
is at the very core of what gives animals
and plants their uniqueness
 New discoveries in genetic techniques
include:
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Copying DNA (PCR)
DNA revealing owner’s id (DNA Profiling)
Mapping DNA (Human Genome Project)
Cutting and pasting genes (gene transfer)
Cloning cells and animals
 These
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
new techniques offer new hope for:
Treatment and vaccines
Creating new plants
Free or convicting people
 PCR
is a laboratory technique which takes
very small quantity of DNA and copies all the
nucleic acids in it to make millions of copies
of the DNA



Solves a very simple problem-not enough DNA to
analyze
http://www.dnalc.org/resources/animations/pcr
.html
http://learn.genetics.utah.edu/content/labs/pc
r/
 This
laboratory technique is
used to separate fragments of
DNA in an effort to identify its
origin

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Enzymes are used to chop up
the long filament of DNA into
varying sized fragments
DNA fragments are placed into
small wells (holes) in the gel
which are aligned along one end
The gel is exposed to an electric
current (positive on one side
and negative on the other)
 The


biggest , heaviest, and least charged particles do
not move easily through the gel
Smallest, least massive and most charged
particles pass through the gel to the other side
with little difficulty
 In

effect is that the
the end
The fragments leave a banded pattern of DNA
 http://learn.genetics.utah.edu/content/labs
/gel/
 https://www.msu.edu/~russellr/portfolio/el
ectrophoresis/electrophoresis.html
 The
process of matching an unknown
sample of DNA with a known sample to see
if they correspond is call DNA profiling

AKA DNA fingerprinting
 If,
after separation by gel electrophoresis,
the:

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Pattern of bands formed by two samples of
DNA fragments are identical, it means that
both most certainly came from the same
individual
Patterns are similar, it means that the two
individuals are most probably related
 Applications

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of DNA profiling
Paternity suits
Match DNA to suspect
Determine how closely related species are to
each other
Determine evolution
 Look
at page 102
 1990-started
an international cooperative
venture-Human Genome Project

Goal: sequence the complete human genome
 2003-annouced

they succeeded the project
Scientist are working on deciphering which
sequences represent genes and which do what
 With
genome libraries of genetic diseases,
doctors can find out exactly where to look if
they think one of their patients might
possess a disease-carrying allele
 Other
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Produce new medications, Steps:
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Advantages:
Find beneficial molecules which are produced
naturally in healthy people
Find out which gene controls the synthesis of a
desirable molecule
Copy that gene and use it as instructions to synthesize
the molecule in a laboratory
Distribute the beneficial molecule as a new treatment
By comparing the genetic makeup of populations
around the world, countless details could be
revealed about ancestries and how humans
migrated and mixed their genes with other
populations over time
 The
technique of taking a gene out of one
organism and placing it in another organism
is a genetic engineering procedure called
gene transfer

It is possible to transfer any species genes into
another species makeup
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Example: proteins used by fish to resist the icy
temperature of arctic waters are now produced by the
modified tomatoes to make them more resistant to
cold
Example: corn, genetically engineered to produce
toxins that kill the bugs which attack it (take protein
from Bacillus thuringiensis)
 The
scissors used for cutting base sequences
are called restriction enzymes

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Restriction enzymes called endonucleases find
and recognize a specific sequence of base pairs
along the DNA molecule (4 or 6 sets of base pairs)
The endonucleases cut the DNA at the specific
points

If both the beginning and the end of a gene are cut,
the gene is released and can be removed from the
donor organisms
 For
pasting genes, the enzyme used is called
DNA Ligase

It recognizes the parts of the base sequences
that are supposed to be clicked together, called
sticky ends, and attach them
 http://www.dnalc.org/resources/animations
/restriction.html
 This
is more complex because a host cell is
needed in addition to the cutting and pasting
enzymes. (most common- Escherichia coli)

Some DNA is found in structures called plasmids

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Plasmids are small circles of extra copies of DNA
floating around inside the cell’s cytoplasm
 To copy a gene, it must be glued into a plasmid
Plasmid is removed from the host cell and cut open
using a restriction endonucleases
 To
copy a gene, it must be
glued into a plasmid

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Plasmid is removed from the host
cell and cut open using a
restriction endonucleases
The gene to be copied is placed
inside the open plasmid
 Process called gene splicing
The plasmid is now called a
recombinant plasmid and can be
used as a vector, a tool for
introducing a new gene into an
organism’s genetic makeup
 In
the final step needed for copying (or
cloning) the gene.

The vector is placed inside the host bacterium
and he bacterium is given its ideal conditions to
grow and proliferate.

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Done by putting the bacterium into a vat of nutritious
liquid kept at a warm temperature
Example: Used to make insulin
 Is
one that has had an artificial genetic
change using the techniques of genetic
engineering

One of the main reasons is to be more
competitive in food production
 Transgenic
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Simplest kind of GM food is one in which an
undesirable gene is removed
In some cases, another more desirable gene is
put in its place.
 First
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plants
Example of GM Food
1994-Flavr Savr tomato-genetically modified to
delay the ripening and rotting process
Another tomato-able to grow in high salinity soil
Rice-engineered to produce beta carotene
 Transgenic
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animals
Haemophilia-Factor IX

Using transgenic sheep
 Genetic
engineering raising many social or
ethical questions

Benefits, promises, and hopes for the future

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GM crops will help farmers by improving food
production
GM crops which produce their own pest-control
substances will be beneficial to the environment
because fewer chemical pesticides will be needed
Using GMOs to produce rare proteins for medicines or
vaccines could be, in a long run, less costly and
produce less pollution than synthesizing such proteins
in laboratories
Farmers can be more in control of what crops or
livestock they produce (selective breeding)
The multinational companies who make GM plants
claim that they will enable farmers in developing
nations to help reduce hunger by using pest-resistant
crops or GM plants which require less water
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No one knows the long-term effects of GMO’s in the
wild
There is a danger that the genes could cross species
Bt-crops which produces toxins to kill insects could
be harmful to humans.
The risks of allergies (may be allergic to one but not
another)
Critics are worried that large portions of the human
food supply will be the property of a small number of
corporations
High tech solutions are not necessarily better than
simpler solutions
A proliferation of genetically modified organisms may
lead to a decrease in biodiversity
 Clones

and cloning
The definition of a clone is a group of genetically
identical organisms or a group of cells artificially
derived form a single parent

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In either case, the resulting cells or organisms were
made using laboratory techniques
In farming, clones have been made for decades for
regenerating plant material or by allowing an in-vitro
fertilized egg or divide to make copies of itself
 1996,
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a sheep by name of Dolly was born
First clone whose genetic material did not
originate from an egg cell
This type of cloning is called reproductive cloning
because it makes an entire individual

Steps to create Dolly
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First the original donor sheep to be cloned, a somatic
cell from the udder was collected and cultured. The
nucleus was removed from a cultured cell
An unfertilized egg was collected from another sheep
and its nucleus was removed
Using a zap of electrical current, the egg cell and the
nucleus form the cultured somatic cell were fused
together
The new cell developed in vitro in a similar way to a
zygote and started to form an embryo
The embryo was placed in a womb of a surrogate
mother sheep
Dolly was born, and was presented to the world as a
clone of the original donor sheep
 Cloning

Therapeutic cloning-simply to make copies of
cells
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using undifferentiated cells
Aim is to develop cells which have not yet gone
through the process of differentiation
Since the first technique in this area involved
using embryos, the cells are referred to as
embryonic stem cells (stem cell research)
 Since
therapeutic cloning starts with the
production of human embryos, it raises
fundamental issues of right and wrong

Is it ethically acceptable to generate a new
human embryo for the sole purpose of medical
research
 Major
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breakthroughs (current research):
Growing skin to repair a serious burn
Growing new heart muscle to repair an ailing
heart
Growing new kidney tissue to rebuild a failing
kidney
 Most
researchers and medical professionals
are against the idea of reproductive cloning
in humans