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Introduction
to
Human
Cloning
TIP 2
Group C
CLFS620 Modern Molecular Genetics
Focus Questions
Answer these questions as you view the Power point.
1. What is cloning?
2. What are the 3 types of human cloning?
3. Compare and contrast therapeutic
cloning and reproductive cloning.
4. What are ESC? What are sources of ESC?
5. Give an example of the usage of each of
the 3 types of human cloning.
What is Cloning?
CLONING is the creation of a copy of a
gene or an entire organism using DNA
from an existing individual, so that the
copy has the same genetic makeup as the
original biological entity.
What is Human Cloning?
Human cloning is the
creation of a
genetically identical
copy of an existing, or
previously existing,
human.
Potential uses of cloning
• Replacing organs and other tissues
• Infertility aid
• Treatment of human diseases
• Creating donor sources
• Gene therapy
• Reversing the aging process
Different Types of Human Cloning
DNA Cloning
also called gene
cloning or
recombinant DNA
Therapeutic Reproductive
Cloning
Cloning
also called
embryo cloning
produces copies
of genes or
segments of genes
produces a
cloned embryo to
create embryonic
stem cells
the transfer of
DNA pieces from
one organism to a
self-replicating
genetic element
cloning for the
use of studying
human
development and
treating disease
the type of
cloning that
produces a
genetically identical
human from
another human
DNA Cloning
What is Gene (DNA) Cloning?
(recombinant DNA)
Recombinant DNA
transfer of a DNA
fragment of
interest from one
organism to a
self-replicating genetic
element,
such as a bacterial
plasmid.
The DNA is then
propagated in
the foreign host cell.
Basic Steps of Gene Cloning
1) Restriction enzymes are
used to cut a fragment of
DNA containing the gene
to be cloned.
2) Fragments of the foreign
DNA are inserted into
plasmid vectors cut open
with the same restriction
enzyme or one which
produces a match end.
3) DNA ligase seals the two
DNA strands together to
produce a recombinant
DNA molecule.
Basic Steps of Gene Cloning
4) The vector transports the gene
into a host cell (usually a
bacterium).
5) Within the host cell the vector
multiplies, producing
numerous identical copies not
only of itself but also of the
gene that it carries.
6) When the host cell divides,
copies of the recombinant
DNA molecule are passed to
the progeny and further vector
replication takes place.
7) After a large number of cell
divisions, a colony or clone of
identical host cells is
produced. Each cell in the
clone contains one or more
copies of the recombinant
DNA molecule. 
The gene is cloned.
Uses of DNA Cloning
1. Isolation of a particular gene,
part of a gene or region of a
genome
2. Production of a desired RNA or
protein molecule in large
quantities
3. Increased production efficiency
for commercially made
enzymes and drugs
4. Modification of existing
organisms so that they express
a trait not previously encoded
in the genome
(transformation)
Uses of DNA Cloning
Therapeutic
Cloning
What is Therapeutic Cloning?
• Therapeutic cloning refers to the medical
procedure by which stem cells are
harvested and grown to mature into
transplantable organs or tissues.
• Creates new identical copies of an
organism’s cells using their own DNA
Therapeutic Cloning
Basic Principle
1. Performed by removing
healthy adult cells from a
patient
2. reprogramming the cell’s
nuclei
3. collecting and growing
embryonic stem cell clones
from the resulting blastocyst
4. and inducing these embryonic
stem cell clones to
differentiate into the stem cell
or mature cell types required
for transplantation.
Therapeutic Cloning is also called
Nuclear Transplantation or
Somatic Cell Nuclear Transfer (SCNT)
SCNT Process
Step 1: Remove the nucleus
from an unfertilized egg cell (A)
while using a suction pipette
(B) to hold the egg cell steady
and a glass needle (C) to
remove the cell’s nucleus.
Step 2: Gently push the glass
needle through the tough shell
that surrounds the egg cell.
The glass needle is used to
remove the nucleus from within
the egg.
http://www.pbs.org/wgbh/nova/sciencenow/3209/04-clon-nf.html
SCNT Process
Step 3: The egg cell’s nucleus
(A) has been released outside of
the egg. This nuclear material will
no longer be needed.
What remains is an “enucleated”
egg (B). The enucleated egg
contains certain molecules and
other important factors that will
help to establish embryonic stem
cells.
Step 4: Inject the nucleus (red
arrow) from a donor cell into
the enucleated egg cell by
easing the tip of the glass
needle deep into the
enucleated egg cell and
depositing the donor nucleus.
SCNT Process
Step 5: After completing the
nuclear transfer, the
unfertilized egg cell is
“activated” using a chemical or
electrical treatment that
stimulates cellular division.
The first division results in two
cells (left image), the next
makes four cells, and so on,
producing an embryo.
Step 6: The proliferating cells
form a structure called a
blastocyst within days, which is
roughly the same size as the
egg cell.
Cells taken from the blastocyst
are embryonic stem cells
(ESC).
Embryonic Stem Cells (ESC)
• are not implanted into the
uterus
• are used to study
development
• may be mixed with
chemicals to
help the cells take on
different properties
• ultimately may be able to
introduce these cells into
an adult
(therapeutic cloning)
Embryonic Stem Cells can be cultured in different laboratory
environments to develop into a specific cell type.
Liver cells
Nerve Cells
Cultured
embryonic
stem cells
(developing an ESC line)
Muscle Cells
Different culture
conditions
Different types of
differentiated cells
Embryonic Stem Cells (ESC)
• Embryonic stem cells (ESC) are cells that have yet to
differentiate
• Unspecialized cells
• Under certain conditions, can be induced to become
cells with specific functions (heart muscle cell,
lymphocyte)
• Sources of ESC
1. Adult Stem cells
2. In-vitro fertilization
3. Umbilical Cord Blood
(Core Blood) Stem cells
Adult Stem Cells
“Adult” Stem Cells
• Somewhat differentiated cells
• Can develop into certain tissues, but not necessarily all tissues in the body
For example: Blood stem cells can develop into RBCs, WBCs, but not muscle
cells
• Can be extracted from adults Ex. from bone marrow sample
Does Not
require the
creation of
an embryo
Adult Stem Cells
The bone marrow is the
spongy core found in the
bones and is a source of
adult stem cells.
These stem cells are the
precursor cells responsible
for the formation of the
blood cells (red blood
cells, platelets, and white
blood cells).
You can also find
these same type of
stem cells in the
blood system:
Peripheral Blood
Stem Cells (PBSCs)
Used to treat
leukemia, other
cancers and various
blood disorders
Less invasive than
collecting bone
marrow, but are
sparse!
What is in Vitro Fertilization (IVF)?
• IVF is the process of fertilization by
manually combining an egg and sperm in
a laboratory setting
• Similar process to in-vitro fertilization
(“test-tube baby”), except embryonic
stems cells are not implanted into
mother
In vitro fertilization
Some procedures involved with IVF manually inject the sperm
into the egg, and others simply allow fertilization to occur by
adding the sperm to the egg in the lab setting.
Why use IVF as a source of stem cells?
• According to a survey conducted in 2003, there
are approximately 400,000 unwanted preembryos in the United States.
(Hoffman, D.I., et al. 2003. Cryopreserved embryos in the United States and their
availability for research. Fertility and Sterility 79: 1063-1069.)
• These may no longer be needed for fertility
purposes and remain frozen or could be
destroyed.
• They may be used in therapeutic cloning
(without being implanted into a uterus).
Umbilical Cord Blood Stem Cells
Multipotent stem cell rich blood found in
the umbilical cord has proven useful in
treating the same types of health
problems as those treated using bone
marrow stem cells.
In 2005, there were more than 1,400 cord
blood transplantations in adults, according
to NETCORD, an international network that
coordinates umbilical cord blood banks.
Differences between embryonic stem
cells and adult stem cells:
-ESC can differentiate into any cell type
(totipotent/pluripotent), while adult SC
have already “committed” to a particular
fate (multipotent).
Some Challenges in Research:
-Adult stem cells are often present in
only minute quantities and can
therefore be difficult to isolate and
purify.
-There is also evidence that they may
not have the same capacity to multiply
as embryonic stem cells do.
-They do not have the development
potential of an ESC.
-Finally, adult stem cells may contain
more DNA abnormalities—caused by
sunlight, toxins, and errors in making
more DNA copies during the course of
a lifetime.
-These potential weaknesses might
limit the usefulness of adult stem cells.
Therapeutic Cloning: Potential Application
• Repair a damaged tissue or group of cells that
can't heal itself.
– This might be accomplished by transplanting ESCs
into the damaged area and directing them to grow
new, healthy tissue.
– It may also be possible to coax stem cells already in
the body to work overtime and produce new tissue.
• To learn more about the causes of
disease/improved research capabilities
• Tissue/Organ transplants
• Potential solutions for currently incurable
degenerative diseases, like cancer
• Substitute case for testing new drugs
Reproductive
Cloning
What is Reproductive Cloning?
Making new,
genetically identical
copies of an
organism, by using its
own DNA
Could possibly create
an “identical” twin,
born years later
Process of Reproductive Cloning
http://cmgm.stanford.edu/biochem118/images/Stem%20Cell%20Slides/08%20Cloning%20Procedures.jpg
Reproductive Cloning
• DNA originally from a mature
somatic cell is added into an
empty oocyte (egg).
• Once the egg has developed
into an early-stage embryo
inside a test-tube, it is
implanted into the womb of an
adult female animal.
• A cloned animal does not
always look identical to the
original animal.
What Animals Have Been Cloned?
Mice
Sheep
Cat
Dog
Mule
Pig
Rat
Monkey
Cows
Chickens
Deer
Horse
Ox
Rabbit
Rhesus
Humans???
Ex. Dolly the Sheep (1996)
Efficiency of
Reproductive Cloning
• Reproductive Cloning is a very inefficient use of
technology.
• There are low success rates in the practice of cloning.
• More that 90% of cloning attempts fail to produce viable
offspring. The cloning of Dolly took 276 attempts.
• Also, more than 100 nuclear transfer procedures could
be required to produce one viable clone.
• Some studies show that after 60 cycles of cell division,
stem cells can mutate and lead to cancer.
Source: http://www.ornl.gov/sci/techresources/Human_Genome/elsi/cloning.shtml
Risks of Human Cloning
• Cloning tends to produce more compromised immune
function and higher rates of infection, tumor growth, and
other disorders.
• About 30% of all clones born alive are affected with large
offspring syndrome and other debilitating conditions.
• Clones show premature aging and have shorter life
spans.
• Also, there is no known facts on how cloning could
impact mental development.
Sources: http://www.ornl.gov/sci/techresources/Human_Genome/elsi/cloning.shtml
Ethical Issues of Human Cloning
-- Could allow for “manufactured” children with
desired traits and characteristics
-- Unrealistic expectations of the clone’s
similarity to the cloned individual
-- Could allow for the cloning of a deceased
individual
-- May violate values of individual freedom,
identity, and autonomy
-- Requires destruction of human embryos in a test
tube
Comparing Therapeutic and
Reproductive Cloning
Therapeutic cloning
• Made in the same way as
reproductive cloning
except the embryo is not
implanted in a uterus
• Donor embryos are killed
when stem cells are
harvested from the
embryos
• The stem cells are used to
grow different types of
tissues
Reproductive cloning
• Made in the same way as
reproductive cloning with
the embryo being
implanted into a uterus to
nurture a living individual
• Donor embryos are killed
during the process.
• Cloned individuals have
shorter lifespans and
possible immune system
malfunctions.
Acceptable or Not to the Public?
DNA or Gene Cloning
Acceptable
Therapeutic Cloning
Acceptable with
reservations: Embryos
are destroyed by stem
cell extraction.
Currently not
Acceptable to clone
humans
Reproductive Cloning
Works Cited
Cloning. The National Human Genome Research Institute. [cited 28 Apr 2009]. Available
from: http://www.genome.gov/pfv.cfm?pageID=25020028
Cloning in Focus. Genetic Science Learning Center at the University of
Utah. [cited 26 Apr 2009]. Available from:
http://learn.genetics.utah.edu/units/cloning/
Perspectives. 2003. Center for Genetics and Society. Human Genetics in the Public
Interest. [cited 1 May 2009]. Available from: http://www.genetics-andsociety.org/perspectives/science.html
Reproductive Cloning. Center for Genetics and Society. [cited 26 Apr 2009]. Available
from: http://geneticsandsociety.org/article.php?id=282
Therapeutic Use of Cell Nuclear Replacement: Therapeutic Cloning.
Medical Research Council. [cited 26 Apr 2009]. Available from:
http://www.reproductivecloning.net/therapeutic_cloning.pdf
United States Department of Energy. Office of Science. “Cloning Fact Sheet.” May 2009.
Human Genome Project Information. Available from:
http://www.ornl.gov/sci/techresources/Human_Genome/elsi/cloning.shtml