Cloning (реферат)

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Cloning

Cloning is the process of creating an identical copy of something. In
HYPERLINK «http://en.wikipedia.org/wiki/Biology» \o «Biology» Biology ,
it collectively refers to processes used to create copies of HYPERLINK
«http://en.wikipedia.org/wiki/DNA» \o «DNA» DNA fragments (Molecular
Cloning), HYPERLINK «http://en.wikipedia.org/wiki/Cell_%28biology%29»
\o «Cell (biology)» cells (Cell Cloning), or HYPERLINK
«http://en.wikipedia.org/wiki/Organisms» \o «Organisms» organisms . The
term also encompases situations, whereby organisms reproduce HYPERLINK
«http://en.wikipedia.org/wiki/Asexual_reproduction» \o «Asexual
reproduction» asexually , but in common parlance refers to
intentionally created copies of organisms.

Etymology

The term clone is derived from ????, the HYPERLINK
«http://en.wikipedia.org/wiki/Greek_language» \o «Greek language» Greek
word for «twig», referring to the process, whereby a new plant can be
created from a twig. In HYPERLINK
«http://en.wikipedia.org/wiki/Horticulture» \o «Horticulture»
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». Since the term entered the popular lexicon in a more
general context, the spelling clone has been used exclusively.

Molecular cloning

Molecular cloning refers to the procedure of isolating a defined DNA
sequence and obtaining multiple copies of it HYPERLINK
«http://en.wikipedia.org/wiki/In_vivo» \o «In vivo» in vivo . Cloning
is frequently employed to amplify DNA fragments containing HYPERLINK
«http://en.wikipedia.org/wiki/Genes» \o «Genes» genes , but it can be
used to amplify any DNA sequence such as HYPERLINK
«http://en.wikipedia.org/wiki/Promoters» \o «Promoters» promoters ,
non-coding sequences and randomly fragmented DNA. It is utilised in a
wide array of biological experiments and practical applications such as
large scale protein production. Occasionally, the term cloning is
misleadingly used to refer to the identification of the HYPERLINK
«http://en.wikipedia.org/wiki/Chromosome» \o «Chromosome» chromosomal
location of a gene associated with a particular phenotype of interest,
such as in positional cloning. In practice, localization of the gene to
a chromosome or genomic region does not necessarily enable one to
isolate or amplify the relevant genomic sequence.

In essence, in order to amplify any DNA sequence in a living organism
that sequence must be linked to an HYPERLINK
«http://en.wikipedia.org/wiki/Origin_of_replication» \o «Origin of
replication» origin of replication , a sequence element capable of
directing the propagation of its self and any linked sequence. In
practice, however, a number of other features are desired and a variety
of specialised cloning HYPERLINK
«http://en.wikipedia.org/wiki/Vectors» \o «Vectors» vectors exist that
allow protein expression, tagging, single stranded HYPERLINK
«http://en.wikipedia.org/wiki/RNA» \o «RNA» RNA and DNA production and
a host of other manipulations.

Cloning of any DNA fragment essentially involves four steps:
fragmentation, ligation, transfection, and screening/selection. Although
these steps are invariable among cloning procedures a number of
alternative routes can be selected, these are summarised as a ‘cloning
strategy’.

Initially, the DNA of interest needs to be isolated to provide a
relevant DNA segment of suitable size. Subsequently, a ligation
procedure is employed whereby the amplified fragment is inserted into a
vector. The vector (which is frequently circular) is linearised by means
of restriction enzymes, and incubated with the fragment of interest
under appropriate conditions with an enzyme called HYPERLINK
«http://en.wikipedia.org/wiki/DNA_ligase» \o «DNA ligase» 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 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
HYPERLINK «http://en.wikipedia.org/wiki/Antibiotic» \o «Antibiotic»
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 is required to confirm that cloning was successful. This may be
accomplished by means of PCR, restriction fragment analysis and/or
HYPERLINK «http://en.wikipedia.org/wiki/DNA_sequencing» \o «DNA
sequencing» DNA sequencing .

Cellular cloning

Cloning a cell means to derive a (clonal) 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 HYPERLINK «http://en.wikipedia.org/wiki/Inoculation» \o
«Inoculation» inoculation of the appropriate medium. However, in the
case of cell cultures from higher organisms, cell cloning is an arduous
task as these cells will not readily grow in standard media.

A valuable tissue culture technique used to clone distinct lineages of
cell lines involves the use of cloning rings (cylinders). According to
this technique, a single-cell suspension of cells which have been
exposed to a HYPERLINK «http://en.wikipedia.org/wiki/Mutagen» \o
«Mutagen» mutagenic agent or drug used to drive HYPERLINK
«http://en.wikipedia.org/wiki/Selection» \o «Selection» selection is
plated at high dilution to create isolated colonies; each arising from a
single and potentially clonally distinct cell. At an early growth stage
when colonies consist of only a few of cells, sterile HYPERLINK
«http://en.wikipedia.org/wiki/Polystyrene» \o «Polystyrene» polystyrene
rings (cloning rings), which have been dipped in grease are placed over
an individual colony and a small amount of HYPERLINK
«http://en.wikipedia.org/wiki/Trypsin» \o «Trypsin» trypsin is added.
Cloned cells are collected from inside the ring and transferred to a new
vessel for further growth.

Organism

Organism cloning refers to the procedure of creating a new mutlicellular
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 HYPERLINK «http://en.wikipedia.org/wiki/Vegetative_reproduction»
\o «Vegetative reproduction» vegetative reproduction ) and some
insects.

Horticultural

The term clone is used in HYPERLINK
«http://en.wikipedia.org/wiki/Horticulture» \o «Horticulture»
horticulture to mean all descendants of a single plant, produced by
HYPERLINK «http://en.wikipedia.org/wiki/Vegetative_reproduction» \o
«Vegetative reproduction» vegetative reproduction or HYPERLINK
«http://en.wikipedia.org/wiki/Apomixis» \o «Apomixis» apomixis . Many
horticultural plant HYPERLINK «http://en.wikipedia.org/wiki/Cultivar»
\o «Cultivar» 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 HYPERLINK
«http://en.wikipedia.org/wiki/Grape» \o «Grape» grapes represent
clones that have been propagated for over two millennia. Other examples
are HYPERLINK «http://en.wikipedia.org/wiki/Potato» \o «Potato»
potato and HYPERLINK «http://en.wikipedia.org/wiki/Banana» \o
«Banana» banana . HYPERLINK «http://en.wikipedia.org/wiki/Grafting»
\o «Grafting» Grafting can be regarded as cloning, since all the
shoots and branches coming from the graft are genetically a clone of a
single individual, although the root systems may be genetically genuine
examples of cloning in the broader biological sense, as they create
genetically identical organisms by biological means, but this particular
kind of cloning has not come under HYPERLINK
«http://en.wikipedia.org/wiki/Ethics» \o «Ethics» ethical scrutiny and
is generally treated as an entirely different kind of operation.

Many HYPERLINK «http://en.wikipedia.org/wiki/Tree» \o «Tree» trees ,
HYPERLINK «http://en.wikipedia.org/wiki/Shrub» \o «Shrub» shrubs ,
HYPERLINK «http://en.wikipedia.org/wiki/Vine» \o «Vine» vines ,
HYPERLINK «http://en.wikipedia.org/wiki/Fern» \o «Fern» ferns and
other HYPERLINK «http://en.wikipedia.org/wiki/Perennials» \o
«Perennials» herbaceous perennials form HYPERLINK
«http://en.wikipedia.org/wiki/Clonal_colony» \o «Clonal colony» clonal
colonies . Parts of a large clonal colony often become detached from the
parent, termed HYPERLINK
«http://en.wikipedia.org/wiki/Fragmentation_%28biology%29» \o
«Fragmentation (biology)» fragmentation , to form separate individuals.
Some plants also form HYPERLINK «http://en.wikipedia.org/wiki/Seed» \o
«Seed» seeds asexually, termed HYPERLINK
«http://en.wikipedia.org/wiki/Apomixis» \o «Apomixis» apomixis , e.g.
HYPERLINK «http://en.wikipedia.org/wiki/Dandelion» \o «Dandelion»
dandelion .

Animals

Clonal derivation exists in nature in some animal species and is
referred to as HYPERLINK
«http://en.wikipedia.org/wiki/Parthenogenesis» \o «Parthenogenesis»
parthenogenesis . An example is the «Little HYPERLINK
«http://en.wikipedia.org/wiki/Fire_ant» \o «Fire ant» Fire Ant » (
HYPERLINK
«http://en.wikipedia.org/w/index.php?title=Wasmannia_auropunctata&action
=edit» \o «Wasmannia auropunctata» Wasmannia auropunctata ), which is
native to HYPERLINK «http://en.wikipedia.org/wiki/Central_America» \o
«Central America» Central and HYPERLINK
«http://en.wikipedia.org/wiki/South_America» \o «South America» South
America but has spread throughout many tropical environments.

Therapeutic Cloning

Therapeutic Cloning refers to a procedure which allows the cloning of
specific body parts and organs to be utilised for medical purposes. This
has not yet being realised, but it is the subject of much active
research. Currently, patients subjected to transplantation are
administered immunosuppresant drugs to prevent recognition of the
foreign transplant by their immune system and its subsequent rejection.
The ability to clonally derive organs from the patients’ own cells would
abolish the need for immunosuppressant drugs and would allow the
patients to live a life without the potentially serious side-effects of
immunosuppresant drugs. More importantly, the ability to clonally derive
organs would alleviate the current shortage of transplants and would
possibly reduce waiting times for translants to become available.

Reproductive Cloning

Reproductive cloning is a technology used to generate an animal that has
the same nuclear DNA as another currently or previously existing animal.
HYPERLINK «http://en.wikipedia.org/wiki/Dolly_the_sheep» \o «Dolly the
sheep» Dolly the sheep was created by reproductive cloning technology.
In a process called » HYPERLINK
«http://en.wikipedia.org/wiki/Somatic_cell_nuclear_transfer» \o «Somatic
cell nuclear transfer» somatic cell nuclear transfer » (SCNT),
scientists transfer genetic material from the nucleus of a donor adult
cell to an egg whose nucleus, and thus its genetic material, has been
removed. The reconstructed egg containing the DNA from a donor cell must
be treated with chemicals or electric current in order to stimulate cell
division. Once the cloned embryo reaches a suitable stage, it is
transferred to the uterus of a female host where it continues to develop
until birth.

Dolly or any other animal created using nuclear transfer technology is
not truly an identical clone of the donor animal. Only the clone’s
chromosomal or nuclear DNA is the same as the donor. Some of the clone’s
genetic materials come from the HYPERLINK
«http://en.wikipedia.org/wiki/Mitochondrion» \o «Mitochondrion»
mitochondria in the HYPERLINK
«http://en.wikipedia.org/wiki/Cytoplasm» \o «Cytoplasm» cytoplasm of
the enucleated egg. Mitochondria, which are HYPERLINK
«http://en.wikipedia.org/wiki/Organelle» \o «Organelle» organelles
that serve as power sources to the cell, contain their own short
segments of DNA, although this is only 0.01% of the total DNA. Acquired
mutations in mitochondrial DNA are believed to play an important role in
the growing process.

Mutations also occur with every cell division so no two cells in an
individual are identical. Thus, nuclear transfer clones from different
maternal lineages are not clones in the strictest sense because 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.

Species cloned

The modern cloning techniques involving HYPERLINK
«http://en.wikipedia.org/wiki/Nuclear_transfer» \o «Nuclear transfer»
nuclear transfer have been successfully performed on several species.
Landmark experiments in chronological order:

HYPERLINK «http://en.wikipedia.org/wiki/Tadpole» \o «Tadpole» Tadpole
: (1952) Many scientists questioned whether cloning had actually
occurred and unpublished experiments by other labs were not able to
reproduce the reported results.

HYPERLINK «http://en.wikipedia.org/wiki/Carp» \o «Carp» Carp : (1963)
In HYPERLINK «http://en.wikipedia.org/wiki/China» \o «China» China ,
HYPERLINK «http://en.wikipedia.org/wiki/Embryologist» \o «Embryologist»
embryologist HYPERLINK «http://en.wikipedia.org/wiki/Tong_Dizhou» \o
«Tong Dizhou» Tong Dizhou cloned a fish. He published the findings in
an obscure Chinese science journal which was never translated into
English. HYPERLINK «http://en.wikipedia.org/wiki/» \l «_note-0» \o «»
[1]

HYPERLINK «http://en.wikipedia.org/wiki/Domestic_sheep» \o «Domestic
sheep» Sheep : (1996) From early embryonic cells by Steen Willadsen.
Megan and Morag cloned from differentiated embryonic cells in June 1995
and HYPERLINK «http://en.wikipedia.org/wiki/Dolly_the_sheep» \o «Dolly
the sheep» Dolly the sheep in 1997.

HYPERLINK «http://en.wikipedia.org/wiki/Rhesus_Monkey» \o «Rhesus
Monkey» Rhesus Monkey : HYPERLINK
«http://en.wikipedia.org/wiki/Tetra_%28monkey%29» \o «Tetra (monkey)»
Tetra (female, January 2000) from embryo splitting

HYPERLINK «http://en.wikipedia.org/wiki/Cattle» \o «Cattle» Cattle :
HYPERLINK
«http://en.wikipedia.org/w/index.php?title=Alpha_and_Beta&action=edit»
\o «Alpha and Beta» Alpha and Beta (males, 2001) and (2005) Brazil
HYPERLINK «http://en.wikipedia.org/wiki/» \l «_note-1» \o «» [2]

HYPERLINK «http://en.wikipedia.org/wiki/Cat» \o «Cat» Cat :
HYPERLINK «http://en.wikipedia.org/wiki/CC_%28cat%29» \o «CC (cat)»
CopyCat «CC» (female, late 2001), HYPERLINK
«http://en.wikipedia.org/wiki/Little_Nicky_%28cat%29» \o «Little Nicky
(cat)» Little Nicky , 2004, was the first cat cloned for commercial
reasons

HYPERLINK «http://en.wikipedia.org/wiki/Mule» \o «Mule» Mule :
HYPERLINK «http://en.wikipedia.org/wiki/Idaho_Gem» \o «Idaho Gem» Idaho
Gem , a john mule born HYPERLINK «http://en.wikipedia.org/wiki/2003»
\o «2003» 2003 — HYPERLINK «http://en.wikipedia.org/wiki/May_4» \o
«May 4» 05-04 , was the first horse-family clone.

HYPERLINK «http://en.wikipedia.org/wiki/Horse» \o «Horse» Horse :
HYPERLINK «http://en.wikipedia.org/wiki/Prometea» \o «Prometea»
Prometea , a Haflinger female born HYPERLINK
«http://en.wikipedia.org/wiki/2003» \o «2003» 2003 — HYPERLINK
«http://en.wikipedia.org/wiki/May_28» \o «May 28» 05-28 , was the first
horse clone.

For a complete list see: HYPERLINK
«http://en.wikipedia.org/wiki/List_of_animals_that_have_been_cloned» \o
«List of animals that have been cloned» List of animals that have been
cloned .

Health aspects

The success rate of cloning has been low: HYPERLINK
«http://en.wikipedia.org/wiki/Dolly_the_sheep» \o «Dolly the sheep»
Dolly the sheep was born after 277 eggs were used to create 29 embryos,
which only produced three lambs at birth, only one of which lived,
Dolly. Seventy calves have been created from 9,000 attempts and one
third of them died young; HYPERLINK
«http://en.wikipedia.org/wiki/Prometea» \o «Prometea» Prometea took
328 attempts, and, more recently, Paris Texas was created after 400
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 HYPERLINK
«http://en.wikipedia.org/wiki/Dolly_the_Sheep» \o «Dolly the Sheep»
Dolly the Sheep had accelerated aging. Aging of this type is thought to
be due to the shortening of HYPERLINK
«http://en.wikipedia.org/wiki/Telomere» \o «Telomere» telomeres ,
regions at the tips of HYPERLINK
«http://en.wikipedia.org/wiki/Chromosome» \o «Chromosome» chromosomes
which prevent genetic threads from fraying every time a HYPERLINK
«http://en.wikipedia.org/wiki/Cell_division» \o «Cell division» cell
divides . Over time telomeres get worn down until cell-division is no
longer possible — this is thought to be a cause of aging. However,
subsequent studies showed that, if anything, Dolly’s telomere were
longer than normal. Dolly died in the year of HYPERLINK
«http://en.wikipedia.org/wiki/2003» \o «2003» 2003 . HYPERLINK
«http://en.wikipedia.org/wiki/Ian_Wilmut» \o «Ian Wilmut» Ian Wilmut
said that Dolly’s early death had nothing to do with cloning but with a
respiratory infection common to lambs raised like Dolly.

Consistent with Dolly’s telomeres being longer, analysis of the
telomeres from cloned cows showed that they were also longer. This
suggests clones could live longer life spans although many died young
after excessive growth. Researchers think that this could eventually be
developed to reverse aging in humans, provided that this is based
chiefly on the shortening of telomeres. Although some work has been
performed on telomeres and aging in nuclear transfer clones, the
evidence is at an early stage.

Human cloning

Human cloning is the creation of a HYPERLINK
«http://en.wikipedia.org/wiki/Genetics» \o «Genetics» genetically
identical copy of an existing, or previously existing HYPERLINK
«http://en.wikipedia.org/wiki/Human» \o «Human» human , by growing
cloned HYPERLINK «http://en.wikipedia.org/wiki/Biological_tissue» \o
«Biological tissue» tissue from that individual. The term is generally
used to refer to artificial human cloning; human clones in the form of
HYPERLINK «http://en.wikipedia.org/wiki/Identical_twin» \o «Identical
twin» identical twins are commonplace, with their cloning occurring
during the natural process of reproduction.

Human cloning is amongst the most controversial forms of the practice.
HYPERLINK «http://en.wikipedia.org/wiki/» \l «_note-3» \o «» [4] There
have been numerous demands for all progress in the human cloning field
to be halted. One of the most ethically questionable problems with human
cloning is farming of organs from clones. For example, many believe it
is unethical to use a human clone to save the life of another. In this
scenario, the cloned human would be euthanized so that the vital organs
could be harvested. This process of renewing the body’s organs would
potentially increase the life expectancy of a human by 50 years. 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 human organisms.
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. This practice
is still morally questionable, but arguably less so than the process of
harvesting said organs from a cloned human being.

The cloning described above is HYPERLINK
«http://en.wikipedia.org/wiki/Human_cloning» \o «Human cloning»
reproductive cloning , not to be confused with HYPERLINK
«http://en.wikipedia.org/wiki/Somatic_cell_nuclear_transfer» \o «Somatic
cell nuclear transfer» research cloning in which only parts (such as
an organ) are cloned using genetic material from a patient’s tissues.

Ethical issues of human cloning

Although the practice of cloning organisms has been widespread for
several thousands of years in the form of horticultural cloning, the
recent technological advancements that have allowed for cloning of
animals (and potentially humans) have been highly controversial. Many
religious groups oppose all forms of cloning, including the potentially
life-saving cloning of individual organs, on the grounds that life
begins at conception. Concerns also exist regarding the protection of
the identity of the individual and the right to protect one’s genetic
identity. In addition, if technology eventually does allow for
successful cloning of humans, prejudice may develop against clones, as
if they were a «lesser» form of human being. Such prejudice could force
clones into a kind of slavery or caste system. The possible social
implications of an artificial human production scheme were famously
explored in the novel HYPERLINK
«http://en.wikipedia.org/wiki/Brave_New_World» \o «Brave New World»
Brave New World .

References

Vogel, Gretchen (2000). «In Contrast to Dolly, Cloning Resets Telomere
Clock in Cattle». Science 288: 641.

Pence, Gregory E. (1998). Who’s Afraid of Human Cloning?. Rowman &
Littlefield.

Heidi B. Perlman. «Scientists Close on Extinct Cloning», Associated
Press, 2000-10-08.

Pence, Gregory E. (2005). Cloning After Dolly: Who’s Still Afraid?

Holloway, Grant. «Cloning to revive extinct species», CNN.com,
2002-05-28.

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