Cloning: For The Cloning of Human Beings, See - For Other Uses, See
Cloning: For The Cloning of Human Beings, See - For Other Uses, See
Cloning: For The Cloning of Human Beings, See - For Other Uses, See
The term clone is derived from κλῶνος, the Greek word for "trunk, branch", referring to
the process whereby a new plant can be created from a twig. In horticulture, the spelling
clon was used until the twentieth century; the final e came into use to indicate the vowel
is a "long o" instead of a "short o".[1][2] Since the term entered the popular lexicon in a
more general context, the spelling clone has been used exclusively.
Contents
[hide]
• 1 Molecular cloning
• 2 Cellular cloning
o 2.1 Unicellular organisms
o 2.2 Cloning in stem cell research
• 3 Organism cloning
o 3.1 Horticultural
o 3.2 Parthenogenesis
o 3.3 Artificial cloning of organisms
3.3.1 Methods
3.3.2 Dolly the Sheep
3.3.3 Water buffalo
3.3.4 Species cloned
3.3.5 Human cloning
3.3.6 Ethical issues of cloning
3.3.7 Cloning extinct and endangered species
• 4 In science fiction
• 5 See also
• 6 References
• 7 External links
Molecular cloning
Main article: Molecular cloning
Although these steps are invariable among cloning procedures a number of alternative
routes can be selected, these are summarized as a 'cloning strategy'.
Initially, the DNA of interest needs to be isolated to provide a DNA segment of suitable
size. Subsequently, a ligation procedure is used where the amplified fragment is inserted
into a vector (piece of DNA). The vector (which is frequently circular) is linearised using
restriction enzymes, and incubated with the fragment of interest under appropriate
conditions with an enzyme called DNA ligase. Following ligation the vector with the
insert of interest is transfected into cells. A number of alternative techniques are
available, such as chemical sensitivation of cells, electroporation, optical injection and
biolistics. Finally, the transfected cells are cultured. As the aforementioned procedures
are of particularly low efficiency, there is a need to identify the cells that have been
successfully transfected with the vector construct containing the desired insertion
sequence in the required orientation. Modern cloning vectors include selectable antibiotic
resistance markers, which allow only cells in which the vector has been transfected, to
grow. Additionally, the cloning vectors may contain colour selection markers, which
provide blue/white screening (??-factor complementation) on X-gal medium.
Nevertheless, these selection steps do not absolutely guarantee that the DNA insert is
present in the cells obtained. Further investigation of the resulting colonies must be
required to confirm that cloning was successful. This may be accomplished by means of
PCR, restriction fragment analysis and/or DNA sequencing.
Cellular cloning
Unicellular organisms
Cloning a cell means to derive a population of cells from a single cell. In the case of
unicellular organisms such as bacteria and yeast, this process is remarkably simple and
essentially only requires the inoculation of the appropriate medium. However, in the case
of cell cultures from multi-cellular organisms, cell cloning is an arduous task as these
cells will not readily grow in standard media.
A useful tissue culture technique used to clone distinct lineages of cell lines involves the
use of cloning rings (cylinders).[4] According to this technique, a single-cell suspension of
cells that have been exposed to a mutagenic agent or drug used to drive selection is plated
at high dilution to create isolated colonies; each arising from a single and potentially
clonal distinct cell. At an early growth stage when colonies consist of only a few of cells,
sterile polystyrene rings (cloning rings), which have been dipped in grease are placed
over an individual colony and a small amount of trypsin is added. Cloned cells are
collected from inside the ring and transferred to a new vessel for further growth.
Somatic cell nuclear transfer, known as SCNT, can also be used to create embryos for
research or therapeutic purposes. The most likely purpose for this is to produce embryos
for use in stem cell research. This process is also called "research cloning" or "therapeutic
cloning." The goal is not to create cloned human beings (called "reproductive cloning"),
but rather to harvest stem cells that can be used to study human development and to
potentially treat disease. While a clonal human blastocyst has been created, stem cell
lines are yet to be isolated from a clonal source.[5]
Organism cloning
Further information: Asexual reproduction
Organism cloning (also called reproductive cloning) refers to the procedure of creating a
new multicellular organism, genetically identical to another. In essence this form of
cloning is an asexual method of reproduction, where fertilization or inter-gamete contact
does not take place. Asexual reproduction is a naturally occurring phenomenon in many
species, including most plants (see vegetative reproduction) and some insects. Scientists
have made some major achievements with cloning, including the asexual reproduction of
sheep and cows. There is a lot of ethical debate over whether or not cloning should be
used. However, cloning, or asexual propagation,[6] has been common practice in the
horticultural world for hundreds of years.
Horticultural
The term clone is used in horticulture to refer to descendants of a single plant which were
produced by vegetative reproduction or apomixis. Many horticultural plant cultivars are
clones, having been derived from a single individual, multiplied by some process other
than sexual reproduction. As an example, some European cultivars of grapes represent
clones that have been propagated for over two millennia. Other examples are potato and
banana and grapes. Grafting can be regarded as cloning, since all the shoots and branches
coming from the graft are genetically a clone of a single individual, but this particular
kind of cloning has not come under ethical scrutiny and is generally treated as an entirely
different kind of operation.
Many trees, shrubs, vines, ferns and other herbaceous perennials form clonal colonies.
Parts of a large clonal colony often become detached from the parent, termed
fragmentation, to form separate individuals. Some plants also form seeds asexually,
termed apomixis, e.g. dandelion.
Parthenogenesis
Methods
Reproductive cloning generally uses "somatic cell nuclear transfer" (SCNT) to create
animals that are genetically identical. This process entails the transfer of a nucleus from a
donor adult cell (somatic cell) to an egg that has no nucleus. If the egg begins to divide
normally it is transferred into the uterus of the surrogate mother. Such clones are not
strictly identical since the somatic cells may contain mutations in their nuclear DNA.
Additionally, the mitochondria in the cytoplasm also contains DNA and during SCNT
this DNA is wholly from the donor egg, thus the mitochondrial genome is not the same as
that of the nucleus donor cell from which it was produced. This may have important
implications for cross-species nuclear transfer in which nuclear-mitochondrial
incompatibilities may lead to death.
Artificial embryo splitting or embryo twinning may also be used as a method of cloning,
where an embryo is split in the maturation before embryo transfer. It is optimally
performed at the 6- to 8-cell stage, where it can be used as an expansion of IVF to
increase the number of available embryos.[7] If both embryos are successful, it gives rise
to monozygotic (identical) twins.
Dolly was publicly significant because the effort showed that the genetic material from a
specific adult cell, programmed to express only a distinct subset of its genes, can be
reprogrammed to grow an entirely new organism. Before this demonstration, it had been
shown by John Gurdon that nuclei from differentiated cells could give rise to an entire
organism after transplantation into an enucleated egg.[9] However, this concept was not
yet demonstrated in a mamallian system.
Cloning Dolly the sheep had a low success rate per fertilized egg; she was born after 237
eggs were used to create 29 embryos, which only produced three lambs at birth, only one
of which lived. Seventy calves have been created and one third of them died young;
Prometea took 277 attempts. Notably, although the first clones were frogs, no adult
cloned frog has yet been produced from a somatic adult nucleus donor cell.
There were early claims that Dolly the Sheep had pathologies resembling accelerated
aging. Scientists speculated that Dolly's death in 2003 was related to the shortening of
telomeres, DNA-protein complexes that protect the end of linear chromosomes.
However, other researchers, including Ian Wilmut who led the team that successfully
cloned Dolly, argue that Dolly's early death due to respiratory infection was unrelated to
deficiencies with the cloning process.
Water buffalo
On September 15, 2007, the Philippines announced its development of Southeast Asia’s
first cloned water buffalo. The Philippine Council for Agriculture, Forestry and Natural
Resources Research and Development (PCARRD), under the Department of Science and
Technology in Los Baños, Laguna approved this project.[10]
Species cloned
The modern cloning techniques involving nuclear transfer have been successfully
performed on several species. Landmark experiments[clarification needed] in chronological order:
• Horse: Prometea, a Haflinger female born 28 May 2003, was the first horse clone.
[23]
• Water Buffalo: Samrupa was the first cloned water buffalo. It was born on
February 6, 2009, at India's Karnal National Diary Research Institute but died five
days later due to lung infection.[24]
• Pyrenean Ibex (2009) was the first extinct animal (extinct 2000) to be cloned back
to life; the clone lived for seven minutes before dying of lung defects.[25]
• Camel: (2009) Injaz, is the first cloned camel.[26]
Human cloning
The various forms of human cloning are controversial.[27] There have been numerous
demands for all progress in the human cloning field to be halted. Most scientific,
governmental and religious organizations oppose reproductive cloning. The American
Association for the Advancement of Science (AAAS) and other scientific organizations
have made public statements suggesting that human reproductive cloning be banned until
safety issues are resolved.[28] Serious ethical concerns have been raised by the future
possibility of harvesting organs from clones.[citation needed] Some people have considered the
idea of growing organs separately from a human organism - in doing this, a new organ
supply could be established without the moral implications of harvesting them from
humans. Research is also being done on the idea of growing organs that are biologically
acceptable to the human body inside of other organisms, such as pigs or cows, then
transplanting them to humans, a form of xenotransplantation.
The first hybrid human clone was created in November 1998, by Advanced Cell
Technologies.[29] It was created from a man's leg cell, and a cow's egg whose DNA was
removed. It was destroyed after 12 days. Since a normal embryo implants at 14 days, Dr
Robert Lanza, ACT's director of tissue engineering, told the Daily Mail newspaper that
the embryo could not be seen as a person before 14 days. While making an embryo,
which may have resulted in a complete human had it been allowed to come to term,
according to ACT: "[ACT's] aim was 'therapeutic cloning' not 'reproductive cloning'"
On January, 2008, Wood and Andrew French, Stemagen's chief scientific officer in
California, announced that they successfully created the first 5 mature human embryos
using DNA from adult skin cells, aiming to provide a source of viable embryonic stem
cells. Dr. Samuel Wood and a colleague donated skin cells, and DNA from those cells
was transferred to human eggs. It is not clear if the embryos produced would have been
capable of further development, but Dr. Wood stated that if that were possible, using the
technology for reproductive cloning would be both unethical and illegal. The 5 cloned
embryos, created in Stemagen Corporation lab, in La Jolla, were destroyed.[30]
Gregory Stock is a scientist and outspoken critic against restrictions on cloning research.
[31]
Bioethicist Gregory Pence also attacks the idea of criminalizing attempts to clone
humans.[citation needed]
The social implications of an artificial human production scheme were famously explored
in Aldous Huxley's novel Brave New World.
On December 28, 2006, the U.S. Food and Drug Administration (FDA) approved the
consumption of meat and other products from cloned animals.[32] Cloned-animal products
were said to be virtually indistinguishable from the non-cloned animals. Furthermore,
companies would not be required to provide labels informing the consumer that the meat
comes from a cloned animal.[33]
Critics have raised objections to the FDA's approval of cloned-animal products for
human consumption, arguing that the FDA's research was inadequate, inappropriately
limited, and of questionable scientific validity.[34][35][36] Several consumer-advocate groups
are working to encourage a tracking program that would allow consumers to become
more aware of cloned-animal products within their food.[37]
Joseph Mendelson, legal director of the Center for Food Safety, said that cloned food still
should be labeled since safety and ethical issues about it remain questionable.
Carol Tucker Foreman, director of food policy at the Consumer Federation of America,
stated that FDA does not consider the fact that the results of some studies revealed that
cloned animals have increased rates of mortality and deformity at birth.
Cloning, or more precisely, the reconstruction of functional DNA from extinct species
has, for decades, been a dream of some scientists. The possible implications of this were
dramatized in the best-selling novel by Michael Crichton and high budget Hollywood
thriller Jurassic Park. In real life, one of the most anticipated targets for cloning was
once the Woolly Mammoth, but attempts to extract DNA from frozen mammoths have
been unsuccessful, though a joint Russo-Japanese team is currently working toward this
goal. And in January 2011, it was reported by Yomiuri Shimbun that a team of scientists
headed by Akira Iritani of Kyoto University had built upon research by Dr. Wakayama,
saying that they will extract DNA from a mammoth carcass that had been preserved in a
Russian laboratory and insert it into the egg cells of an African elephant in hopes of
producing a mammoth embryo. The researchers said they hoped to produce a baby
mammoth within six years.[38]
In 2001, a cow named Bessie gave birth to a cloned Asian gaur, an endangered species,
but the calf died after two days. In 2003, a banteng was successfully cloned, followed by
three African wildcats from a thawed frozen embryo. These successes provided hope that
similar techniques (using surrogate mothers of another species) might be used to clone
extinct species. Anticipating this possibility, tissue samples from the last bucardo
(Pyrenean Ibex) were frozen in liquid nitrogen immediately after it died in 2000.
Researchers are also considering cloning endangered species such as the giant panda,
ocelot, and cheetah. The "Frozen Zoo" at the San Diego Zoo now stores frozen tissue
from the world's rarest and most endangered species.[39][40]
In 2002, geneticists at the Australian Museum announced that they had replicated DNA
of the Thylacine (Tasmanian Tiger), extinct about 65 years previous, using polymerase
chain reaction.[41] However, on February 15, 2005 the museum announced that it was
stopping the project after tests showed the specimens' DNA had been too badly degraded
by the (ethanol) preservative. On 15 May 2005 it was announced that the Thylacine
project would be revived, with new participation from researchers in New South Wales
and Victoria.
In January 2009, for the first time, an extinct animal, the Pyrenean ibex mentioned above
was cloned, at the Centre of Food Technology and Research of Aragon, using the
preserved DNA of the skin samples from 2001 and domestic goat egg-cells. (The ibex
died shortly after birth due to physical defects in its lungs.) [42] One of the continuing
obstacles in the attempt to clone extinct species is the need for nearly perfect DNA.
Cloning from a single specimen could not create a viable breeding population in sexually
reproducing animals. Furthermore, even if males and females were to be cloned, the
question would remain open whether they would be viable at all in the absence of parents
that could teach or show them their natural behavior.
Vertebrate cloning poses little risk to the environment, but it can consume scarce
conservation resources, and its chances of success in preserving species seem poor. To
date, the conservation benefits of transgenics and vertebrate cloning remain entirely
theoretical, but many of the risks are known and documented. Conservation biologists
should devote their research and energies to the established methods of conservation,
none of which require transgenics or vertebrate cloning.[43]
In science fiction
Cloning has been used in countless science fiction works throughout the years. Human
cloning is usually the most popular kind that is used in these particular works, mainly due
to fact that it brings up controversial questions of identity.[44] For example, in C.J.
Cherryh’s novel Cyteen, and in Aldous Huxley’s novel Brave New World, human
cloning is a major plot device that not only drives the story along but also makes the
reader think critically about what identity means.
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