The possibility of human cloning, raised when Scottish scientists at
Roslin Institute created the much-celebrated sheep "Dolly" (Nature
385, 810-13, 1997), aroused worldwide interest and concern because
of its scientific and ethical implications. The feat, cited by Science
magazine as the breakthrough of 1997, also generated uncertainty over
the meaning of "cloning" --an umbrella term traditionally used by scientists
to describe different processes for duplicating biological material.
What is cloning? Are there different types of cloning?
When the media report on cloning in the news, they are usually talking
about only one type called reproductive cloning. There are different types
of cloning however, and cloning technologies can be used for other purposes
besides producing the genetic twin of another organism. A basic understanding
of the different types of cloning is key to taking an informed stance
on current public policy issues and making the best possible personal
decisions. The following three types of cloning technologies will be discussed:
(1) recombinant DNA technology or DNA cloning, (2) reproductive cloning,
and (3) therapeutic cloning.
Recombinant DNA Technology or DNA Cloning
The terms "recombinant DNA technology," "DNA cloning,"
"molecular cloning,"or "gene cloning" all refer to
the same process: the transfer of a DNA fragment of interest from one
organism to a self-replicating genetic element such as a bacterial
The DNA of interest can then be propagated in a foreign host cell. This
technology has been around since the 1970s, and it has become a common
practice in molecular biology labs today.
studying a particular gene often use bacterial plasmids to generate multiple
copies of the same gene. Plasmids are self-replicating extra-chromosomal
circular DNA molecules, distinct from the normal bacterial genome (see
image to the right). Plasmids and other types of cloning vectors are used
by Human Genome Project researchers to copy genes and other pieces of
chromosomes to generate enough identical material for further study.
To "clone a gene," a DNA fragment containing the gene of interest
is isolated from chromosomal DNA using restriction enzymes and then united with
a plasmid that has been cut with the same restriction enzymes. When the fragment
of chromosomal DNA is joined with its cloning vector in the lab, it is called
a "recombinant DNA molecule." Following introduction into suitable
host cells, the recombinant DNA can then be reproduced along with the host cell
Plasmids can carry up to 20,000 bp of foreign DNA. Besides bacterial
plasmids, some other cloning vectors include viruses, bacteria artificial
chromosomes (BACs), and yeast artificial chromosomes (YACs). Cosmids are
artificially constructed cloning vectors that carry up to 45 kb of foreign
DNA and can be packaged in lambda phage particles for infection into E.
coli cells. BACs utilize the naturally occurring F-factor plasmid
found in E. coli to carry 100 to 300 kb DNA inserts. A YAC is
a functional chromosome derived from yeast that can carry up to 1 MB of
foreign DNA. Bacteria are most often used as the host cells for recombinant
DNA molecules, but yeast and mammalian cells also are used.
Sheep Has Died at Age 6
Dolly, the first mammal to be cloned from adult DNA, was put down
by lethal injection Feb. 14, 2003. Prior to her death, Dolly had
been suffering from lung cancer and crippling arthritis. Although
most Finn Dorset sheep live to be 11 to 12 years of age, postmortem
examination of Dolly seemed to indicate that, other than her cancer
and arthritis, she appeared to be quite normal. The unnamed sheep
from which Dolly was cloned had died several years prior to her
creation. Dolly was a mother to six lambs, bred the old-fashioned
Reproductive cloning is a technology used to generate an animal that has the
same nuclear DNA as another currently or previously existing animal. Dolly
was created by reproductive cloning technology. In a process called "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 mitochondria in the cytoplasm of the enucleated egg. Mitochondria,
which are organelles that serve as power sources to the cell, contain
their own short segments of DNA. Acquired mutations in mitochondrial DNA
are believed to play an important role in the aging process.
Dolly's success is truly remarkable because it proved that the genetic
material from a specialized adult cell, such as an udder cell programmed
to express only those genes needed by udder cells, could be reprogrammed
to generate an entire new organism. Before this demonstration, scientists
believed that once a cell became specialized as a liver, heart, udder,
bone, or any other type of cell, the change was permanent and other unneeded
genes in the cell would become inactive. Some scientists believe that
errors or incompleteness in the reprogramming process cause the high rates
of death, deformity, and disability observed among animal clones.
Therapeutic cloning, also called "embryo cloning," is the production
of human embryos for use in research. The goal of this process is not
to create cloned human beings, but rather to harvest stem cells that can
be used to study human development and to treat disease. Stem cells are
important to biomedical researchers because they can be used to generate
virtually any type of specialized cell in the human body. Stem cells are
extracted from the egg after it has divided for 5 days. The egg at this
stage of development is called a blastocyst. The extraction process destroys
the embryo, which raises a variety of ethical concerns. Many researchers
hope that one day stem cells can be used to serve as replacement cells
to treat heart disease, Alzheimer's, cancer, and other diseases. See more
on the potential use of cloning in organ transplants.
In November 2001, scientists from Advanced Cell Technologies (ACT), a
biotechnology company in Massachusetts, announced that they had cloned
the first human embryos for the purpose of advancing therapeutic research.
To do this, they collected eggs from women's ovaries and then removed
the genetic material from these eggs with a needle less than 2/10,000th
of an inch wide. A skin cell was inserted inside the enucleated egg to
serve as a new nucleus. The egg began to divide after it was stimulated
with a chemical called ionomycin. The results were limited in success.
Although this process was carried out with eight eggs, only three began
dividing, and only one was able to divide into six cells before stopping.
How can cloning technologies be used?
Recombinant DNA technology is important for learning about other related
technologies, such as gene therapy, genetic engineering of organisms,
and sequencing genomes. Gene therapy can be used to treat certain genetic
conditions by introducing virus vectors that carry corrected copies of
faulty genes into the cells of a host organism. Genes from different organisms
that improve taste and nutritional value or provide resistance to particular
types of disease can be used to genetically engineer food crops. See Genetically
Modified Foods and Organisms for more information. With genome sequencing,
fragments of chromosomal DNA must be inserted into different cloning vectors
to generate fragments of an appropriate size for sequencing. See
a diagram on constructing clones for sequencing.
If the low success rates can be improved (Dolly was only one success out of
276 tries), reproductive cloning can be used to develop efficient ways to reliably
reproduce animals with special qualities. For example, drug-producing animals
or animals that have been genetically altered to serve as models for studying
human disease could be mass-produced.
Reproductive cloning also could be used to repopulate endangered animals or
animals that are difficult to breed. In 2001, the first clone of an endangered
wild animal was born, a wild ox called a gaur. The young gaur died from
an infection about 48 hours after its birth. In 2001, scientists in Italy
reported the successful cloning of a healthy baby mouflon, an endangered
wild sheep. The cloned mouflon is living at a wildlife center in Sardinia.
Other endangered species that are potential candidates for cloning include
the African bongo antelope, the Sumatran tiger, and the giant panda. Cloning
extinct animals presents a much greater challenge to scientists because
the egg and the surrogate needed to create the cloned embryo would be
of a species different from the clone.
Therapeutic cloning technology may some day be used in humans to produce
whole organs from single cells or to produce healthy cells that can replace
damaged cells in degenerative diseases such as Alzheimer's or Parkinson's.
Much work still needs to be done before therapeutic cloning can become
a realistic option for the treatment of disorders.
What animals have been cloned?
Scientists have been cloning animals for many years. In 1952, the first
animal, a tadpole, was cloned. Before the creation of Dolly, the first
mammal cloned from the cell of an adult animal, clones were created from
embryonic cells. Since Dolly, researchers have cloned a number of large
and small animals including sheep, goats, cows, mice, pigs, cats, rabbits,
and a gaur. See Cloned Animals below. All these
clones were created using nuclear transfer technology.
Hundreds of cloned animals exist today, but the number of different species
is limited. Attempts at cloning certain species such as monkeys, chickens,
horses, and dogs, have been unsuccessful. Some species may be more resistant
to somatic cell nuclear transfer than others. The process of stripping
the nucleus from an egg cell and replacing it with the nucleus of a donor
cell is a traumatic one, and improvements in cloning technologies may
be needed before many species can be cloned successfully.
Can organs be cloned for use in transplants?
Scientists hope that one day therapeutic cloning can be used to generate
tissues and organs for transplants. To do this, DNA would be extracted
from the person in need of a transplant and inserted into an enucleated
egg. After the egg containing the patient's DNA starts to divide, embryonic
stem cells that can be transformed into any type of tissue would be harvested.
The stem cells would be used to generate an organ or tissue that is a
genetic match to the recipient. In theory, the cloned organ could then
be transplanted into the patient without the risk of tissue rejection.
If organs could be generated from cloned human embryos, the need for organ
donation could be significantly reduced.
Many challenges must be overcome before "cloned organ" transplants
become reality. More effective technologies for creating human embryos,
harvesting stem cells, and producing organs from stem cells would have
to be developed. In 2001, scientists with the biotechnology company Advanced
Cell Technology (ACT) reported that they had cloned the first human embryos;
however, the only embryo to survive the cloning process stopped developing
after dividing into six cells. In February 2002, scientists with the same
biotech company reported that they had successfully transplanted kidney-like
organs into cows. The team of researchers created a cloned cow embryo
by removing the DNA from an egg cell and then injecting the DNA from the
skin cell of the donor cow's ear. Since little is known about manipulating
embryonic stem cells from cows, the scientists let the cloned embryos
develop into fetuses. The scientists then harvested fetal tissue from
the clones and transplanted it into the donor cow. In the three months
of observation following the transplant, no sign of immune rejection was
observed in the transplant recipient.
Another potential application of cloning to organ transplants is the
creation of genetically modified pigs from which organs suitable for human
transplants could be harvested . The transplant of organs and tissues
from animals to humans is called xenotransplantation.
Why pigs? Primates would be a closer match genetically to humans, but
they are more difficult to clone and have a much lower rate of reproduction.
Of the animal species that have been cloned successfully, pig tissues
and organs are more similar to those of humans. To create a "knock-out"
pig, scientists must inactivate the genes that cause the human immune
system to reject an implanted pig organ. The genes are knocked out in
individual cells, which are then used to create clones from which organs
can be harvested. In 2002, a British biotechnology company reported that
it was the first to produce "double knock-out" pigs that have
been genetically engineered to lack both copies of a gene involved in
transplant rejection. More research is needed to study the transplantation
of organs from "knock-out" pigs to other animals.
What are the risks of cloning?
Reproductive cloning is expensive and highly inefficient. More than 90%
of cloning attempts fail to produce viable offspring. More than 100 nuclear
transfer procedures could be required to produce one viable clone. In
addition to low success rates, cloned animals tend to have more compromised
immune function and higher rates of infection, tumor growth, and other
disorders. Japanese studies have shown that cloned mice live in poor health
and die early. About a third of the cloned calves born alive have died
young, and many of them were abnormally large. Many cloned animals have
not lived long enough to generate good data about how clones age. Appearing
healthy at a young age unfortunately is not a good indicator of long term
survival. Clones have been known to die mysteriously. For example, Australia's
first cloned sheep appeared healthy and energetic on the day she died,
and the results from her autopsy failed to determine a cause of death.
In 2002, researchers at the Whitehead Institute for Biomedical Research
in Cambridge, Massachusetts, reported that the genomes of cloned mice
are compromised. In analyzing more than 10,000 liver and placenta cells
of cloned mice, they discovered that about 4% of genes function abnormally.
The abnormalities do not arise from mutations in the genes but from changes
in the normal activation or expression of certain genes.
Problems also may result from programming errors in the genetic material
from a donor cell. When an embryo is created from the union of a sperm
and an egg, the embryo receives copies of most genes from both parents.
A process called "imprinting" chemically marks the DNA from
the mother and father so that only one copy of a gene (either the maternal
or paternal gene) is turned on. Defects in the genetic imprint of DNA
from a single donor cell may lead to some of the developmental abnormalities
of cloned embryos.
For more details on the risks associated with cloning, see the Cloning
Problems links below.
Should humans be cloned?
Physicians from the American Medical Association and scientists with the American
Association for the Advancement of Science have issued formal public statements
advising against human reproductive cloning. Currently, the U.S. Congress is
considering the passage of legislation that could ban human cloning. See the
Policy and Legislation links below.
Due to the inefficiency of animal cloning (only about 1 or 2 viable offspring
for every 100 experiments) and the lack of understanding about reproductive
cloning, many scientists and physicians strongly believe that it would
be unethical to attempt to clone humans. Not only do most attempts to
clone mammals fail, about 30% of clones born alive are affected with "large
offspring syndrome" and other debilitating conditions. Several cloned
animals have died prematurely from infections and other complications.
The same problems would be expected in human cloning. In addition, scientists
do not know how cloning could impact mental development. While factors
such as intellect and mood may not be as important for a cow or a mouse,
they are crucial for the development of healthy humans. With so many unknowns
concerning reproductive cloning, the attempt to clone humans at this time
is considered potentially dangerous and ethically irresponsible. See the
Cloning Ethics links below for more information
about the human cloning debate.
(most resources focus on reproductive cloning)
Human Beings (PDF) - Report and Recommendations of the National
Bioethics Advisory Commission – Published in 1997. Also available
in HTML from the National Bioethics Advisory Commission Publications