CLONING
Lecture Notes for Biotechnology
What is Cloning?
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To most people, the term “cloning”
means making a copy of an individual.
In biology, cloning can have different
uses depending on what is being
copied.
Cloning Molecules and Cells
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DNA makes a copy when it replicates to
make two molecules
A cell divides by mitosis to make two
identical daughter cells.
Cloning Organisms
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Botanists and home gardeners have been
cloning plants for centuries – take a “cutting”
or piece of a plant and put in soil and it will
form a new plant.
Animals that reproduce by asexual
reproduction, such as small female freshwater
crustaceans, produce offspring which are
genetic copies of the mother
Cloning Vertebrate Animals
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In the 1960’s, John Gurdon
experimented with a technique called
nuclear transplantation
He destroyed the nuclei of unfertilized
eggs of the African clawed toad with UV
light and replaced them with nuclei
taken from intestinal cells of tadpoles of
the same species
Gurdon (cont)
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A small percentage of the eggs with the
transplanted nuclei developed past the
cleavage stage into tadpoles and adults
Later experiments using the
transplanted nuclei from adult toad
skin, kidney, heart and lung cells
produced the same results
Cloning Mammals
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In 1997 Ian Wilmut and colleagues in
Scotland announced the birth of “Dolly” the
first cloned mammal.
In a procedure similar to Gurdon’s, called
somatic cell nuclear transfer, the lab group
removed the nuclei from cells in the
mammary gland of a sheep and placed them
in enucleated eggs of another sheep.
This link shows images of somatic cell nuclear
transfer
(http://www.advancedcell.com/scnt.htm)
Dolly (cont)
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The eggs were stimulated to begin
dividing by treating them with either
chemicals or electricity
Some of the eggs starting cleaving and
were placed in the uteruses of other
sheep. Only one attempt of 277 was
successful, producing Dolly.
Dolly (cont)
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Dolly developed lung cancer and
arthritis and was euthanized in 2003 at
the age of 6 years.
Most sheep of Dolly’s breed live to 11 or
12 years.
Other Cloned Mammals
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Since Dolly, other mammal species have been
successfully cloned.
These include cow, goat, cat, pig, mule and
gaur.
In many of these attempts, such as the one
producing CC the kitten, the donor nucleus is
not removed from its cell but the donor cell is
fused with an enucleated egg.
CC the Kitten
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Click on the link below to see Copy Cat
(CC) the cloned kitten (Nature, 2002)
CC has a different coat coloration, so
she is not identical to the nucleus donor
http://www.nature.com/nature/journal/
v415/n6874/fig_tab/nature723_F1.html
CC the Kitten
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CC is result of the fusion of a donor
cumulus cell and a recipient egg
She is the sole live birth of 188 nuclear
transfer procedures. 82 produced
blastocysts.
Cloning Human Cells
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Researchers distinguish between
therapeutic and reproductive cloning
In therapeutic cloning, the cloned egg is
allowed to divide for a few days and
then the cells are separated from each
other and saved as stem cells for
potential use to treat diseases
Cloning Human Cells
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In 2001 a Worcester, MA company
called, ACT, announced the first human
cloning. Of eight cloned eggs, only one
made it to the six-cell stage
Later, a ban was put on this type of
research by the US government if it was
supported by public funds
Cloning Human Cells
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Research continues in other countries
and privately-funded labs in the US
Recently scientists in South Korea
announced they successfully cloned a
human embryo to the blastocyst stage
and then separated the cells to begin
stem cell lines
Cloning Human Cells
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In reproductive cloning, the cloned egg
would be allowed to divide the the
blastocyst stage and the the embryo
would be implanted into a woman’s
uterus to continue development to birth
Cloning Human Cells
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A few years ago, a group called the
Raelians, announced the birth of a
cloned human baby
They offered no scientific evidence to
support their claim and it was dismissed
by the scientific community
Are Clones Identical Copies?
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As shown by CC the kitten, clones may
not be identical to the nuclear donor
The uterine environment has an
influence on development
Another reason may be the source of
the mitochondria for the cloned embryo
Are Clones Identical Copies?
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Mitochondria have their own circular strand of
DNA coding for genes involved with their
structure and function.
Mitochondrial genetic mutations cause several
genetic diseases
Mitochondria are usually only maternally
derived, the sperm does not supply any to a
zygote.
Are Clones Identical Copies?
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In strict nuclear transfer, the
mitochondria would be supplied by the
recipient cell not the nuclear DNA
donor.
In the cell fusion technique, the
mitochondria would come from both
cells.
Potential Uses of Cloning
Technology
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Gene therapy
Genetic engineering of organisms
Sequencing genomes
Reproductive cloning of animals to
produce some with special qualities
Other Potential Uses
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Reproductive cloning of animals to
repopulate endangered species. This
has already been done with the gaur, a
wild ox, and a mouflon, a wild sheep.
Therapeutic cloning to produce whole
organs for transplants in humans
Therapeutic cloning to produce healthy
cells to replace diseased cells
Risks of Cloning
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The technique rarely works and is very
expensive
Cloned animals, like Dolly, do not live
long and have a variety of aliments
A third of cloned calves born alive die
young and many are abnormally large
Risks of Cloning
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In cloned mice, 4% of the genes
function abnormally
Genetic imprinting of sperm-derived vs
egg-derived genes is not possible
References
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http://www.ornl.gov/sci/techresources/
Human_Genome/elsi/cloning.shtml
Prentice, D. A. 2003. Stem Cells and
Cloning. Benjamin Cummings
Cell biology: A cat cloned by nuclear
transplantation. 2002. Nature. 415:859