Animal Cloning

ANIMAL CLONING
The production of exact copies of individuals identical to one of their parents is the called animal
clening. It produces the same type of animals and hence it is also known as animal xeroxing, the
animals thus produced are called clone or xerox animals.
Clone animals are not at all considered as offspring. This is because these individuals contain
chromosomes from only one parent. They are simply copies of an animal. Therefore, cloning is
almost equivalent to vegetative propagation of plants. The first animal clone was a frog cloned
by Thomas J. King and Robert W. Briggs in 1952.
In the modern context, cloning is the transfer of a diploid nucleus of a somatic cell into an
enucleated egg to produce clone identical to the nucleus donor. All individuals of a clone there-
fore contain nuclear genome of one parent and mitochondrial genome of the other parent (egg
donor).
Cloning is done by adopting two techniques.
;They are nuclear transfer and embryonic stem cells method
Animal cloning would help to mankind in following ways:

1. Keeping the copies of pet animals in the memories of those pets.
2. Production of identical individuals for genetics research.
3. Preservation of rare and endangered species on the earth.
4. Production of identical individuals with desired sex.
5. Restoration of best traits in living form
I. Nuclear Transfer
The transfer of diploid somatic nuclea into enucleated egg cells is known as nuclear transfer or
nuclear transplantation. The individual donating the diploid nucleus is called do nor and the
individual giving egg cell is called recipient. Nuclear transfer is done with micro injection
method or electrofusion method.
Nuclear transfer helps for cloning in animals such as frog. Amoeba, Xenopus, Drosophila, sheep,
cow, dog, cat, deer, man, etc
. 1. Cloning in Frogs
T.J. King and R.W. Briggs first produced cloned tadpoles of frog in 1952. They performed
nuclear transplantation in Rana pipiens to pro duce cloned tadpoles. The frog cloning involved
the following steps:
1. Blastula stage embryos were collected from the breeding ground of R. pipiens.
2. the blastocysts were taken in a test tube and treated with trypsin to separate the cells of the
blastocysts.
3. The cells were dropped on a glass slide and mounted on the microscope stage. 4. A
micropipette was inserted into the cell and nucleus therein was drawn in.
5. The nucleus was then kept in an isotonic solution. Thus many nuclei were obtained and
preserved in the solution.
6. A mature egg was taken from a female frog by dissecting its body.
7. The egg was activated by pricking it with a glass needle. The egg nucleus therefore moved
towards the animal pole.
8. The egg nucleus was carefully removed by injecting a micropipette while viewing the egg
under a microscope. Thus enucleated eggs were prepared.
9. The diploid nucleus of blastocysts was then injected into enucleated egg using a micropipette
10. The micropipette was drawn out carefully and the injected site was gently forced with two
small needles to stop oozing of cytoplasm from the egg.
11. After nuclear transplantation, the egg was maintained in water contained in a small bottle.
The egg underwent cleavage, blastulation and gastrulation and became a tadpole. In this way
about 80-90% of transplanted eggs gave rise to tadpoles.
In 19625, J.B. Gurden transferred diploid nucleus taken from intestinal cells of Xenopus laevis
into enucleated eggs. The nucleus transplanted eggs developed into tadpoles of Xeno- pus and
then grew into frogs. If the donor nucleus was damaged during handling, the development of the
egg was found to be abnormal. His early experiments therefore gave poor results: only 2-5% of
nuclear transplants alone developed into swimming tadpoles.
This research however proves that diploid nucleus of mature somatic cells can behave as a
zygote nuclens when it is injected into enleated egg cells.
Methodology of Cow Cloning
Scientists in Japan have successfully cloned a cow in December 1998. The methodology of cow
cloning is briefly given below:
1. Some somatic cells are taken from udder and kept in a balanced salt solution.
2. The somatic cells in the solution are incubated at cold temperatures below 5°C for about a
week.
3. Another cow or the same cow is treated with gonadotropin injection to stimulate egg
development. As a result more eggs are produced
4. Mature eggs are taken from the cow using a laparoscope and placed in a nutrient solution.
5. The haploid nucleus in the egg is sucked out using a micropipette under a microscope. This
process is known as enucleation. The egg cell without nucleus is called enucleate egg.
6. The somatic cell is exposed to room temperature for 2 hrs and is then fused with the
enucleated egg cell.
7. The fused cell is grown in a nutrient medium till it has attained 8-16 cells stage
8. The globular mass of cells (embryo) is then implanted into the uterus of the cow.
9. The cow produces a calf clone in 270 days.
The calf is a clone of cow that has contributed somatic nucleus. Cow cloning is one of the latest
developments in cattle breeding and development
Uses
Cow cloning methodology has the following advantages:
1. Production of Female Calves: Since somatic cells are taken from udder, the calves are all
female. They are identical with the parent contributing the somatic cell. Cow cloning thus
increases the number of milking cows.
2. Production of High Quality Calves from Inferior Quality Cows: Holstein- Freecian variety
cows produce a large quantity of milk but they suffer with frequent abortion. Malui breed cow is
poor in milk yield but resistant to abortion.
Somatic cells of Holstein-Freecian cow art fused with enucleated egg cell of Malui breed The
fused cell is cultured and introduced int uterus of the Malui breed cow. The Malui cov produces a
calve which is the duplicate copy c Holstein-Freecian cow. Thus cow cloning help us to breed
high quality cows.
1. Somatic Cells from Dead Breed Somatic cells can also be taken from dead ca tle for
cloning to produce clones of dead cattle
. Embryonic Stem Cell Method

The undifferentiated cells of embryos that can give rise to specialized cells via differentiation are
called embryonic stem cells (ESCs). The ECSs can be isolated from blastcoyst stage embryos.
These ECSs are used in animal cloning in cases where the nucleus transplanted egg proceeds
differentiation prior to first cleavage. Embryonic stem cells are used in nuclear transplantation as
well as in chimera production.
Embryonic stem cells are considered for animal cloning because of the following reasons:
1. Stem cells can be maintained in cell cultures by using irradiated fibroblasts as a feeder layer.
2. They have retained the embryo forming property even after prolonged duration of sub-
cultures.
3. They maintain stable karyotypes in cultures.
4. They can be propagated continuously without differentiation.
5. Stem cells can be used as nucleus donors.
6. They are stably maintained in embryos when the cells are transfused into the embryos.
7. Their genome can be modified suitably for genetic manipulation.
Production of Chimera
A chimera is a mix of two or more animals. It has the characteristics of two or more organisms
which contributed cells to make up the body of that chimera. Chimeras are produced by taking
cells from two carly embryos (blastocysts) and mixing them together. Chimera is also known as
mixed animal. It is an offspring of four parents.
Chimera can be made by embryonic stem cell transfer, embryo fusion and injection of embryonic
carcinoma cell.
1. Embryonic Stem Cell Transfer
In this method, embryonic stem cells taken from one embryo are transplanted into another
embryo and then the transplanted embryo is grown into a chimera. This method is practised in
mice and man. This method involves the following steps
1. Eggs are isolated from a black mouse (donor) and are fertilized in vitro with semen. collected
from a male.
2. The fertilized eggs are grown in a nutrient solution to grow them into young embryos as the
stage of blastocysts.
3.It is performed to mix the traits of two individuals in one organism for breeding pur- poses.
. Embryo Fusion
It is nothing but the fusion of parts of two young embryos of animals to produce one offspring.
It is used to create chimeric mice. It was first practised by Beatrice Mintz at the Cancer Research
Institute, Philadelphia in 1960. In this experiment, young embryos (blasto-cysts) are taken from
two strains of mice. A male and female of each strain is allowed to mate for producing young
embryos. The embryos taken from the two strains are transferred to a microscopic field; the
outer membrane of the embryos is removed carefully using microneedles. Then the embryos of
the two strains are mixed together to bring out the fu sion of the embryos into large mass of cells.
The large mass of cells develop a membrane around it to become a blastocyst. Thus a few large
blas tocysts are made in culture medium.
A surrogate mother is mated with a sterile male to bring her for proper stage of implanta tion.
Then the blastocysts are implanted in her uterus to rear the blastocysts into mice siplings. The
surrogate mother will produce chimeric mice siplings when the pregnancy period is over.
Several chimeras have been created by adopting embryo fusion technique. Geep chimera of goat
and sheep. Humouse is a chimera of man and mouse. Teratomous cells in embryos are called
embryonic carcinoma
3. Injection of Embryonic Carcinoma cells (ECC)
cells (ECC). They are totipotent enough to grow into complete organisms. When ECC taken
from an embryo is injected into another embryo and then the embryo is implanted into a
surrogate mother, the surro- mouse gives birth to chimeric mice siplings. method has been used
to engineer embryos producing transgenic chimeric mice. Chimera production has been strongly
opposed by several people as it is against the God's rules of creation. Hence, chimeras have not
been publicized among the people. Even scientists have not taken attempts to create chimeras of
higher mammals and man.

Animal Cloning

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ANIMAL CLONING

Animal cloning would help to mankind in following ways:

. Embryonic Stem Cell Method

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