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Chapter 8

The moral issues posed by human cloning are profound and have implications for today and for future generations. Today's overwhelming and bipartisan House action to prohibit human cloning is a strong ethical statement, which I commend. We must advance the promise and cause of science, but must do so in a way that honors and respects life.

President George W. Bush, July 2001

We must not say to millions of sick or injured human beings "go ahead and die and stay paralyzed because we believe a clump of cells is more important than you are."

Representative Jerrold Nadler (D-NY), July 2001

The Human Genome Project defines three distinct types of cloning. The first is the use of highly specialized deoxyribonucleic acid (DNA) technology to produce multiple, exact copies of a single gene or other segment of DNA to obtain sufficient material to examine for research purposes. This process produces cloned collections of DNA known as clone libraries. The second kind of cloning involves the natural process of cell division to create identical copies of the entire cell. These copies are called a cell line. The third type of cloning, reproductive cloning, is the one that has received the most attention in the mass media. This is the process that generates complete, genetically identical organisms such as Dolly, the famous Scottish sheep cloned in 1996 and named after the entertainer Dolly Parton.

Cloning may also be described by the technology used to perform it. For example, the term recombinant DNA technology describes the technology and mechanism of DNA cloning. Also known as molecular cloning, or gene cloning, it involves the transfer of a specific DNA fragment of interest to researchers from one organism to a self-replicating genetic element of another species such as a bacterial plasmid. (See Figure 8.1.) The DNA under study may then be reproduced in a host cell. This technology has been in use since the 1970s and is a standard practice in molecular biology laboratories.

Just as GenBank is an online public repository of the human genome sequence, the Clone Registry database is a sort of "public library." Used by genome sequencing centers to record which clones have been selected for sequencing, which sequencing efforts are currently under way, and which are finished and represented by sequence entries in GenBank, the Clone Registry may be freely accessed by scientists worldwide. To effectively coordinate all of this information, a standardized system of naming clones is essential. The nomenclature used is shown in Figure 8.2.


Molecular cloning is performed to enable researchers to have many copies of genetic material available in the laboratory for the purpose of experimentation. Cloned genes allow researchers to examine encoded proteins and are used to sequence DNA. Gene cloning also allows researchers to isolate and experiment on the genes of an organism. This is particularly important in terms of human research; in instances where direct experimentation on humans might be dangerous or unethical, experimentation on cloned genes is often practical and feasible.

Cloned genes are also used to produce pharmaceutical drugs, insulin, clotting factors, human growth hormone, and industrial enzymes. Before the widespread use of molecular cloning, these proteins were difficult and expensive to manufacture. For example, before recombinant DNA technology, insulin (a pancreatic hormone that regulates blood glucose levels) used by people with diabetes was extracted and purified from cow and pig pancreases. Because the amino acid sequences of insulin from cows and pigs are slightly different from those in human insulin, some patients experienced adverse immune reactions to the nonhuman "foreign insulin." The recombinant human version of insulin is identical to human insulin so it does not produce an immune reaction.

Figure 8.3 shows how a gene is cloned. First, a DNA fragment containing the gene being studied is isolated from chromosomal DNA using restriction enzymes. It is joined with a plasmid (a small ring of DNA found in many bacteria that can carry foreign DNA) that has been cut with the same restriction enzymes. When the fragment of chromosomal DNA is joined with its cloning vector (cloning vectors, such as plasmids and yeast artificial chromosomes, introduce foreign DNA into host cells), it is called a recombinant DNA molecule. Once it has entered into the host cells, the recombinant DNA can be reproduced along with the host cell DNA.

Another molecular cloning technique that is simpler and less expensive than the recombinant cloning method is the polymerase chain reaction (PCR). PCR has also been dubbed "molecular photocopying" because it amplifies DNA without the use of a plasmid. Figure 6.5 in chapter 6 shows how PCR is used to generate a virtually unlimited number of copies of a piece of DNA.


Another way to describe and classify cloning is by its purpose. Organismal or reproductive cloning is a technology used to produce a genetically identical organisman animal with the same nuclear DNA as an existing, or even an extinct, animal.

The reproductive cloning technology used to create animals is called somatic cell nuclear transfer (SCNT). In SCNT scientists transfer genetic material from the nucleus of a donor adult cell to an enucleated egg (an egg from which the nucleus has been removed). This eliminates the need for fertilization of an egg by a sperm. The reconstructed egg containing the DNA from a donor cell is treated with chemicals or electric current to stimulate cell division. Once the cloned embryo reaches a suitable stage, it is transferred to the uterus of a surrogate (female host), where it continues to grow and develop until birth. Figure 8.4 shows the entire SCNT process that culminates in the transfer of the embryo into the surrogate mother.

Organisms or animals generated using SCNT are not perfect or identical clones of the donor organism or "parent" animal. Although the clone's nuclear DNA is identical to the donor's, some of the clone's genetic materials come from the mitochondria in the cytoplasm of the enucleated egg. Mitochondria, the organelles that serve as energy sources for the cell, contain their own short segments of DNA called mtDNA. Acquired mutations in the mDNA contribute to differences between clones and their donors and are believed to influence the aging process.

Dolly the Sheep Paves the Way for Other Cloned Animals

In 1952 scientists transferred a cell from a frog embryo into an unfertilized egg, which then developed into a tadpole. This process became the prototype for cloning. Ever since, scientists have been cloning animals. The first mammals were also cloned from embryonic cells in the 1980s. In 1997 cloning became headline news when, following more than 250 failed attempts, Ian Wilmut and his colleagues at the Roslin Institute in Edinburgh, Scotland, successfully cloned a sheep, which they named Dolly. Dolly was the first mammal cloned from the cell of an adult animal, and since then researchers have used cells from adult animals and various modifications of nuclear transfer technology to clone a range of animals, including sheep, goats, cows, mice, pigs, cats, and rabbits.

To create Dolly, the Roslin Institute researchers transplanted a nucleus from a mammary gland cell of a Finn Dorsett sheep into the enucleated egg of a Scottish blackface ewe and used electricity to stimulate cell division. The newly formed cell divided and was placed in the uterus of a blackface ewe to gestate. Born several months later, Dolly was a true clonegenetically identical to the Finn Dorsett mammary cells and not to the blackface ewe, which served as her surrogate mother. Her birth revolutionized the world's understanding of molecular biology, ignited worldwide discussion about the morality of generating new life through cloning, prompted legislation in dozens of countries, and launched an ongoing political debate in the U.S. Congress.

Dolly was the object of intense media and public fascination. She proved to be a basically healthy clone and produced six lambs of her own through normal sexual means. Before her death by lethal injection on February 14, 2003, Dolly had been suffering from lung cancer and arthritis. An autopsy (postmortem examination) of Dolly revealed that, other than her cancer and arthritis, she was anatomically like other sheep.

In February 1997 Don Wolf and his colleagues at the Oregon Regional Primate Center in Beaverton successfully cloned two rhesus monkeys using laboratory techniques that had previously produced frogs, cows, and mice. It was the first time that researchers used a nuclear transplant to generate monkeys. The monkeys were created using different donor blastocysts (early-stage embryos), so they were not clones of one anothereach monkey was a clone of the original blastocyst that had developed from a fertilized egg. Unfortunately, neither of the cloned monkeys survived past the embryonic stage.

An important distinction between the process that created Dolly and the one that produced the monkeys was that unspecialized embryonic cells were used to create the monkeys, whereas a specialized adult cell was used to create Dolly. The Oregon experiment was followed closely in the scientific and lay communities because, in terms of evolutionary biology and genetics, primates are closely related to humans. Researchers and the public speculated that if monkeys could be cloned, it might become feasible to clone humans.

In May 2001 BresaGen Limited, an Australian biotechnology firm, announced the birth of that country's first cloned pig. The pig was cloned from cells that had been frozen in liquid nitrogen for more than two years, and the company used technology that was different from the process used to clone Dolly the sheep. The most immediate benefit of this new technology was to improve livestockcloning enables breeders to take some animals with superior genetics and rapidly produce more. Biomedical scientists were especially attentive to this research because of its potential for xenotransplantationthe use of animal organs for transplantation into humans. Pig organs genetically modified so that they are not rejected by the human immune system could prove to be a boon to medical transplantation.

During the same year the first cat was cloned, and the following year rabbits were successfully cloned. In January 2003 researchers at Texas A&M University reported that cloned pigs behaved normallyas expected for a litter of pigsbut were not identical to the animals from which they were cloned in terms of food preferences, temperament, and how they spent their time. The researchers explained the variation as arising from the environment and epigenetic (not involving DNA sequence change) factors, causing the DNA to line up differently in the clones. Epigenetic activity is defined as any gene-regulating action that does not involve changes to the DNA code and that persists through one or more generations, and it may explain why abnormalities such as fetal death occur more frequently in cloned species.

On May 4, 2003, a cloned mulethe first successful clone of any member of the horse familywas born in Idaho. The clone was not just any mule, but the brother of the world's second fastest racing mule. Named Idaho Gem, the cloned mule was created by researchers at the University of Idaho and Utah State University. The researchers attributed their success to changes in the culture medium they used to nurture the eggs and embryos.

In August 2003 scientists at the Laboratory of Reproductive Technology in Cremona, Italy, were the first to clone a horse. The Italian scientists, Cesare Galli et al., describe their cloning technique in "Pregnancy: A Cloned Horse Born to Its Dam Twin" (Nature, August 7, 2003).

The mule was cloned from cells extracted from a mule fetus, whereas the cloned horse's DNA came from her adult mother's skin cells. There were other differences as well. The University of Idaho and Utah State University researchers harvested fertile eggs from mares, removed the nucleus of each egg, and inserted DNA from cells of a mule fetus. The reconstructed eggs were then surgically implanted into the wombs of female horses. In contrast, the Italian scientists harvested hundreds of eggs from mare carcasses, cultured the eggs, removed their DNA, and replaced it with DNA taken from either adult male or female horse skin cells.

In May 2004 the first bull was cloned from a previously cloned bull in a process known as serial somatic cell cloning or recloning. Before the bull, the only other successful recloning efforts involved mice. Chikara Kubota, X. Cindy Tian, and Xiangzhong Yang, the successful research team, describe their techniques in "Serial Bull Cloning by Somatic Cell Nuclear Transfer" (Nature Biotechnology, May 23, 2004). Their effort was also cited in the Guinness Book of World Records as the "largest clone in the world."

At the close of 2004 a South Korean research team reported cloning macaque monkey embryos, which would be used as a source of stem cells. Conservationists then focused research efforts on cloning rare and endangered species. In April 2005 Texas A&M University announced the first successfully cloned foal in the United States. That same month, Korean scientists at Seoul National University (SNU) cloned a dog they dubbed "Snuppy." In May 2005 the Brazilian Agricultural Research Corporation, Embrapa, reported the creation of two cloned calves from a Junquiera cow, which is an endangered species.

Cloning Endangered Species

Reproductive cloning technology may be used to repopulate endangered species such as the African bongo antelope, the Sumatran tiger, and the giant panda, or animals that reproduce poorly in zoos or are difficult to breed. On January 8, 2001, scientists at Advanced Cell Technology (ACT), a biotechnology company in Massachusetts, announced the birth of the first clone of an endangered animal, a baby bull gaura large wild ox from India and Southeast Asianamed Noah. Noah was cloned using the nuclei of frozen skin cells taken from an adult male gaur that had died eight years earlier. The skin cell nuclei were joined with enucleated cow eggs, one of which was implanted into a surrogate cow mother. Unfortunately, the cloned gaur died from an infection within days of its birth. The same year scientists in Italy successfully cloned an endangered wild sheep. Cloning an endangered animal is different from cloning a more common animal because cloned animals need surrogate mothers to be carried to term. The transfer of embryos is risky, and researchers are reluctant to put an endangered animal through the rigors of surrogate motherhood, opting to use nonendangered domesticated animals whenever possible.

Cloning extinct animals is even more challenging than cloning living animals because the egg and the surrogate mother used to create and harbor the cloned embryo are not the same species as the clone. Furthermore, for most already extinct animal species such as the woolly mammoth or dinosaur, there is insufficient intact cellular and genetic material from which to generate clones. In the future, carefully preserving intact cellular material of imperiled species may allow for their preservation and propagation.

In April 2003 ACT announced the birth of a healthy clone of a Javan banteng, an endangered cattlelike animal native to Asian jungles. The clone was created from a single skin cell, taken from another banteng before it died in 1980, which had remained frozen until it was used to create the clone. The banteng embryo gestated in a standard beef cow in Iowa.

Born April 1, 2003, the cloned banteng developed normally, growing its characteristic horns and reaching an adult weight of about 1,800 pounds. He was nicknamed Stockings and, as of 2007, lived at the San Diego Zoo. Hunting and habitat destruction have reduced the number of banteng, which once lived in large numbers in the bamboo forests of Asia, by more than 75% from 1983 to 2003.

In August 2005 the Audubon Nature Institute in New Orleans, Louisiana, reported that two unrelated endangered African wildcat clones had given birth to eight babies. Their births confirmed that clones of wild animals can breed naturally, which is vitally important for protecting endangered animals on the brink of extinction.

Reproductive Human Cloning

In December 2002 a religious sect known as the Raelians made news when their private biotechnology firm, Clonaid, announced that they had successfully delivered "the world's first cloned baby." The announcement, which could not be independently verified or substantiated, generated unprecedented media coverage and was condemned in the scientific and lay communities. At least some of the media frenzy resulted from the beliefs of the Raeliansnamely, the sect contends that humans were created by extraterrestrial beings. In 2005 Clonaid claimed to have produced at least thirteen cloned children, but as of 2007 had not yet offered any proof of their existence.

Clonaid's announcement brought attention to the fact that several laboratories around the world had embarked on clandestine efforts to clone a human embryo. For example, in 2002 a U.S. fertility specialist, Panayiotis Zavos, claimed to be collaborating with about two dozen international researchers to produce human clones. Another doctor focusing on fertility issues, Severino Antinori, attracted media attention when he maintained that hundreds of infertile couples in Italy and thousands in the United States had already enrolled in his human cloning initiative. Neither these researchers nor anyone else had offered proof of successful reproductive human cloning as of early 2007.


Therapeutic cloning (also called embryo cloning) is the creation of embryos for use in biomedical research. The objective of therapeutic cloning is not to create clones but to obtain stem cells. Stem cells are "master cells" capable of differentiating into multiple other cell types. This potential is important to biomedical researchers because stem cells may be used to generate any type of specialized cell, such as nerve, muscle, blood, or brain cells. Many scientists believe that stem cells can not only provide a ready supply of replacement tissue but also may hold the key to developing more effective treatments for common disorders such as heart disease and cancer as well as degenerative diseases such as Alzheimer's and Parkinson's. Researchers believe that in the future it may be possible to induce stem cells to grow into complete organs.

Advocates of therapeutic cloning point to other treatment benefits such as using stem cells to generate bone marrow for transplants. They contend that scientists could use therapeutic cloning to manufacture perfectly matched bone marrow using the patient's own skin or other cells. This would eliminate the problem of rejection of foreign tissue associated with bone marrow transplant and other organ transplantation. Stem cells also have the potential to repair and restore damaged heart and nerve tissue. Furthermore, there is mounting evidence to suggest that stem cells from cloned embryos have greater potential as medical treatments than stem cells harvested from unused embryos at fertility clinics, which are created by in vitro fertilization and are now the major source of stem cells for research. These prospective benefits are among the most compelling arguments in favor of cloning to obtain embryonic stem cells.

Stem cells used in research are harvested from the blastocyst after it has divided for five days, during the earliest stage of embryonic development. Many people regard human embryos as human beings or at least potential human beings and consider their destruction, or even using techniques to obtain stem cells that might imperil their future viability, as immoral or unethical.

In November 2001 the ACT researchers Jose B. Cibelli et al. reported in "Somatic Cell Nuclear Transfer in Humans: Pronuclear and Early Embryonic Development" (e-biomed: The Journal of Regenerative Medicine, November 26, 2001) that they had created a cloned human embryo, and, unlike groups that had claimed to have done this before, they published their results. The ACT press release "Advanced Cell Technology, Inc. (ACT) Today Announced Publication of Its Research on Human Somatic Cell Nuclear Transfer and Parthenogenesis" (November 25, 2001, boasted that this achievement offered "the first proof that reprogrammed human cells can supply tissue" and asserted that this accomplishment was a vital first step toward the objective of therapeutic cloningusing cloned embryos to harvest embryonic stem cells able to grow into replacement tissue perfectly matched to individual patients. To clone the human embryos, Cibelli et al. collected women's eggs and painstakingly removed the genetic material from the eggs with a thin needle. A skin cell was inserted inside each of eight enucleated eggs, which were then chemically stimulated to divide. Just three of the eight eggs began dividing, and only one reached six cells before cell division ceased.

That same year investigators at the South Australian Research and Development Institute used lambs to experiment with therapeutic cloning. The goal was to replace cells stricken with Parkinson's disease with healthy ones derived from a cloned embryo. In 2003 researchers in Italy reported successfully using adult stem cells to cure mice that had a form of multiple sclerosis. The scientists injected the diseased mice with stem cells that had been extracted from the brains of adult mice reproduced in the laboratory. Postmortem examination of the mice showed that the stem cells had migrated to and then repaired damaged areas of the nerves and brain.

In August 2003 a Chinese research team led by Huizhen Sheng, an American-trained scientist working at the Shanghai Second Medical University, reported that it had made human embryonic stem cells by combining human skin cells with rabbit eggs. Their accomplishment was published in the Chinese scientific journal Cell Research, a peer-reviewed publication of the Shanghai Institute of Cell Biology and the Chinese Academy of Sciences. The researchers removed the rabbit eggs' DNA and injected human skin cells inside them. The eggs then grew to form embryos containing human genetic material. After several days the embryos were dissected to extract their stem cells.

In February 2004 scientists at Seoul National University in South Korea reported in the journal Science that they had successfully cloned healthy human embryos, removed embryonic stem cells, and grown them in mice. In January 2006, following a lengthy investigation, Seoul National University concluded that the research reported in Science had been fabricated. As a result, the journal retracted the article along with another study by the same author. In May 2006, the investigator, Hwang Woo-suk, was charged with fraud, embezzlement, and violating South Korea's bioethics statutes.

In 2005 Wilmut was granted a license by the British government to clone human embryos to generate stem cell lines to study motor neuron disease (MND). Wilmut and his colleagues are working to clone embryos to generate stem cells that would in turn become motor neurons with MND-causing gene defects. By observing the stem cells grow into neurons, the researchers hope to discover what causes the cells to degenerate. Their research involves comparing the stem cells with healthy and diseased cells from MND patients to gain a better understanding of the illness and to test potential drug treatments.

Human reproductive cloning remains illegal in Britain but therapeutic cloningcreating embryos as a source of stem cells to cure diseasesis allowed on an approved basis. The license granted to Wilmut and his colleagues is the second one granted by Britain's Human Fertilisation and Embryology Authority.

In July 2006 the researchers Deepa Deshpande et al. restored movement to paralyzed rats using a new method that demonstrates the potential of embryonic stem cells to restore function to humans suffering from neurological disorders. They published their results in "Recovery from Paralysis in Adult Rats Using Embryonic Stem Cells" (Annals of Neurology, July 2006). Although clinical trials in humans are still years away, the results of this research represent an important advance in the quest for a cure for paralysis and other neurological disorders.

In October 2006 Kevin A. D'Amour et al., in "Production of Pancreatic Hormone-Expressing Endocrine Cells from Human Embryonic Stem Cells" (Nature Biotechnology, October 19, 2006), reported developing a process to turn human embryonic stem cells into pancreatic cells that can produce insulin and other hormones. The researchers anticipate testing these cells in animals in 2008 and if the animal studies are successful, then clinical trials in human patients may begin as soon as 2009.

Three studiesVolker Schächinger et al. in "Intracoronary Bone Marrow-Derived Progenitor Cells in Acute Myocardial Infarction," Ketil Lunde et al. in "Intracoronary Injection of Mononuclear Bone Marrow Cells in Acute Myocardial Infarction," and Birgit Assmus et al. in "Transcoronary Transplantation of Progenitor Cells after Myocardial Infarction"describing the use of stem cells in the treatment of heart disease were published in the September 21, 2006, issue of the New England Journal of Medicine. The studies produced conflicting results: Schächinger and his colleagues reported benefits for patients who had suffered myocardial infarction (heart attack). Lunde and his contributors found no benefit from stem cell treatment of such patients. Assmus and her collaborators studied patients with chronic heart failure, who did show improvement after treatment. In the editorial "Cardiac Cell TherapyMixed Results from Mixed Cells" in the same issue of the journal, Antony Rosenzweig writes that the three studies "provide a realistic perspective on this approach while leaving room for cautious optimism and underscoring the need for further study."

Rick Weiss, in "Stem Cell Work Shows Promise and Risks" (Washington Post, October 23, 2006), reports that research conducted at the University of Rochester Medical Center using nerve cells grown from human embryonic stem cells to treat rats afflicted with Parkinson's disease produced mixed results. The treatment reduced the animals' symptoms, but caused tumors in the rodents' brains. The researchers acknowledged that their work showed both the promise and risks associated with stem cell treatments.

Research Promises Therapeutic Benefits without Cloning

In "Homologous Recombination in Human Embryonic Stem Cells" (Nature Biotechnology, March 2003), Thomas P. Zwaka and James A. Thomson report that they used human embryonic stem cells to splice out individual genes and substituted different genes in their place. Their accomplishment was heralded as a first step toward the goal of regenerating parts of the human body by transplanting either stem cells or tissues grown from stem cells into patients. Zwaka and Thomson used electrical charges and chemicals to make the cells' membranes permeable; the cells allowed the customized genes to enter, and they then found and replaced their counterparts in the cells' DNA.

The ability to make precise genetic changes in human stem cells could be used to boost their therapeutic potential or make them more compatible with patients' immune systems. Some researchers assert that the success of this bioengineering feat might eliminate the need to pursue the hotly debated practice of therapeutic cloning, but others caution that such research could heighten concerns among those who fear that stem cell technology will lead to the creation of "designer babies," which are bred for specific characteristics such as appearance, intelligence, or athletic prowess.

In May 2003 the University of Pennsylvania researcher Hans R. Schöler and his colleagues announced another historic first: The researchers transformed ordinary mouse embryo cells into egg cells in laboratory dishes ("Scientists Produce Mouse Eggs from Embryonic Stem Cells, Demonstrating Totipotency Even In Vitro," ScienceDaily, May 2, 2003). Schöler selected from a population of stem cells the ones that bore certain genetic traits suggesting the potential to become eggs. They then isolated those in laboratory dishes. Eventually, the cells morphed into two kinds of cells, including young egg cells. The eggs matured normally and appeared to be healthy in terms of their appearance, size, and gene expression. When cultured for a few days, the eggs also underwent spontaneous division and formed structures resembling embryos, a process called parthenogenesis. This finding implies that the eggs were fully functional and likely could be fertilized with sperm.

Once refined, this technology could be applied to produce egg cells in the laboratory that would enable scientists to engineer traits into animals and help conservationists rebuild populations of endangered species. It offers researchers the chance to observe mammalian egg cells as they mature, a process that occurs unseen within the ovary. The technology also offers an unparalleled opportunity to learn about meiosis (reduction division), the process of cell division during which an egg or sperm disgorges half of its genes so it can join with a gamete of the opposite sex. There are many potential medical benefits as well. For example, women who cannot make healthy eggs could use this technology to ensure healthy offspring.

Like many new technologies, transforming cells into eggs simultaneously resolves existing ethical issues and creates new ones. For example, because the embryonic stem cells spontaneously transformed themselves into eggs, this procedure overcomes many of the ethical objections to cloning, which involves creating offspring from a single parent. However, it also paves the way for the creation of "designer eggs" from scratch and, if performed with human cells, could redefine the biological definitions of mothers and fathers.

In September 2003 efforts to transform stem cells into sperm were successful. Toshiaki Noce and his colleagues in Tokyo, Japan, observed male mouse embryonic stem cells that developed spontaneously, with some cells actually becoming germ cells. When the researchers transplanted the germ cells into testicular tissue, the cells underwent meiosis and formed sperm cells. One possible medical application of this technology would be to assist couples who are infertile because the male cannot produce healthy sperm. One of the ethical issues that might arise would be the potential for two men to both be biological fathers of a child. Another is the potential to generate a human being who never had any parents using two laboratory-grown stem cells, one transformed into a sperm and the other into an egg. Many ethicists advise consideration of such issues before permitting human experimentation.

In 2004 the National Institutes of Health (NIH) reported that researchers from the University of Pennsylvania School of Veterinary Medicine used cells from mice to grow sperm progenitor cells in a laboratory culture (November 3, 2004, Known as spermatogonial stem cells, the progenitor cells are incapable of fertilizing egg cells but give rise to cells that develop into sperm. The researchers transplanted the cells into infertile mice, which were then able to produce sperm and father offspring that were genetically related to the donor mice.

This breakthrough has many potential applications, including developing new treatments for male infertility and extending the reproductive lives of endangered species. Researchers will also attempt to genetically manipulate the sperm cells grown in a culture medium and then implant the cells into animals. In this way they could introduce new traits into laboratory animals and livestock, such as disease resistance. The culture technique offers researchers additional opportunities to investigate the potential of spermatogonial stem cells as a source for adult stem cells to replace diseased or injured tissue.

New Methods of Obtaining Stem Cells without Destroying Embryos

In "Embryonic and Extraembryonic Stem Cell Lines Derived from Single Mouse Blastomeres" (Nature, January 12, 2006), Young Chung et al. report that embryonic stem cell cultures could be derived from single cells of mouse embryos. Irina Klimanskaya et al., in "Human Embryonic Stem Cell Lines Derived from Single Blastomeres" (Nature, August 23, 2006), describe a technique for removing a single cellcalled a blastomerefrom a three-day-old embryo with eight to ten cells and using a biochemical process to create embryonic stem cells from the blastomere. The method of removing a cell from the embryo is much like the technique used for preimplantation genetic diagnosis, which is performed to screen the cell for genetic defects. The researchers note that human embryonic stem cell lines derived from a single blastomere were comparable to lines derived with conventional techniques. Although Klimanskaya and her colleagues assert that the new method "will make it far more difficult to oppose this research," opponents of stem cell research contend that the new technique is morally unacceptable because even a single cell removed from an early embryo may have the potential to produce a life.

Another technique reported in 2006 can obviate the need for embryonic stem cells. Erika Check notes in "Simple Recipe Gives Adult Cells Embryonic Powers" (Nature, July 6, 2006) that researchers in the United Kingdom discovered the gene, called nanog, that is the key to "reprogramming" adult cells back to an embryonic state. The reprogramming of adult cells using nanog may make it possible for scientists to generate cells that specialize and develop into every type of cell in the body without the controversial use of human embryonic stem cells.


The difficulty and low success rate of much animal reproductive cloning (an average of just one or two viable offspring result from every one hundred attempts) and the as-yet-inadequate understanding about reproductive cloning have prompted many scientists to deem it unethical to attempt to clone humans. Many attempts to clone mammals have failed, and about one-third of clones born alive suffer from anatomical, physiological, or developmental abnormalities that are often debilitating. Some cloned animals have died prematurely from infections and other complications at rates higher than conventionally bred animals, and some researchers anticipate comparable outcomes from human cloning. Furthermore, scientists cannot yet describe or characterize how cloning influences intellectual and emotional development. Even though the attributes of intelligence, temperament, and personality may not be as important for cattle or other primates, they are vital for humans. Without considering the myriad religious, social, and other ethical concerns, the presence of so many unanswered questions about the science of reproductive cloning has prompted many investigators to consider any attempts to clone humans as scientifically irresponsible, unacceptably risky, and morally unallowable.

On August 9, 2001, President George W. Bush ( announced his decision to allow federal funds to be used for research on existing human embryonic stem cell lines as long as the derivation process (which begins with the removal of the inner cell mass from the blastocyst) had already been initiated and the embryo from which the stem cell line was derived no longer had the possibility of development as a human being. The president established the following criteria that research studies must meet to qualify for federal funding:

  • The stem cells must have been drawn from an embryo created for reproductive purposes that was no longer needed for these purposes.
  • Informed consent must have been obtained for the donation of the embryo and no financial inducements provided for donation of the embryo.

In January 2002 the Panel on Scientific and Medical Aspects of Human Cloning was convened by the National Academy of Sciences; the National Academy of Engineering; the Institute of Medicine Committee on Science, Engineering, and Public Policy; and the National Research Council, Division on Earth and Life Studies Board on Life Sciences. Following the panel, the report Scientific and Medical Aspects of Human Cloning (January 2002, was issued that called for a ban on human reproductive cloning.

The panel recommended a legally enforceable ban with substantial penalties as the best way to discourage human reproductive cloning experiments in both the public and private sectors. It cautioned that a voluntary measure might be ineffective because many of the technologies needed to accomplish human reproductive cloning are widely accessible in private clinics and other organizations that are not subject to federal regulations.

The panel did not, however, conclude that the scientific and medical considerations that justify a ban on human reproductive cloning are applicable to nuclear transplantation to produce stem cells. In view of their potential to generate new treatments for life-threatening diseases and advance biomedical knowledge, the panel recommended that biomedical research using nuclear transplantation to produce stem cells be permitted. Finally, the panel encouraged ongoing national discussion and debate about the range of ethical, societal, and religious issues associated with human cloning research.

On February 14, 2002, the American Association for the Advancement of Science (AAAS;, the world's largest general scientific organization, affirmed a legally enforceable ban on reproductive cloning; however, the AAAS supported therapeutic or research cloning using nuclear transplantation methods under appropriate government oversight. Similarly, the American Medical Association (AMA; April 6, 2006,, a national physicians' organization, issued a formal public statement against human reproductive cloning. The AMA statement cautioned that human cloning failures could jeopardize promising science and genetic research and prevent biomedical researchers and patients from realizing the potential benefits of therapeutic cell cloning.

On April 10, 2002, President Bush called on the Senate to back legislation banning all types of human cloning ( In his plea to the Senate, Bush said:

Science has set before us decisions of immense consequence. We can pursue medical research with a clear sense of moral purpose or we can travel without an ethical compass into a world we could live to regret. Science now presses forward the issue of human cloning. How we answer the question of human cloning will place us on one path or the other. Human cloning is deeply troubling to me, and to most Americans. Life is a creation, not a commodity. Our children are gifts to be loved and protected, not products to be designed and manufactured. Allowing cloning would be taking a significant step toward a society in which human beings are grown for spare body parts, and children are engineered to custom specifications; and that's not acceptable. I believe all human cloning is wrong, and both forms of cloning ought to be banned, for the following reasons. First, anything other than a total ban on human cloning would be unethical. Research cloning would contradict the most fundamental principle of medical ethics, that no human life should be exploited or extinguished for the benefit of another.

On September 25, 2002, Elias Zerhouni, the director of the NIH, testified before the Senate Appropriations Subcommittee on Labor, Health and Human Services, and Education in favor of advancing the field of stem cell research ( Zerhouni exhorted Congress to continue to fund both human embryonic stem cell research and adult stem cell research simultaneously to learn as much as possible about the potential of both types of cells to treat human disease. He observed that many studies that do not involve human subjects must be performed before any new therapy is tested on human patients. These preclinical studies include tests of the long-term survival and fate of transplanted cells, as well as tests of the safety, toxicity, and effectiveness of the cells in treating specific diseases in animals. Zerhouni promised that trials using human subjects, the clinical research phase, would begin only after the basic foundation had been established. Despite Zerhouni's impassioned plea and subsequent efforts to advance stem cell research, at the close of 2006 U.S. law continued to ban federal funding of any research that might harm human embryos.

Moral and Ethical Objections to Human Cloning

People who oppose human cloning are as varied as the interests and institutions they support. Religious leaders, scientists, politicians, philosophers, and ethicists argue against the morality and acceptability of human cloning. Nearly all objections hinge, to various degrees, on the definition of human life, beliefs about its sanctity, and the potentially adverse consequences for families and society as a whole.

In an effort to stimulate consideration of and debate about this critical issue, the President's Council on Bioethics examined the principal moral and ethical objections to human cloning in Human Cloning and Human Dignity: An Ethical Inquiry (July 2002, The council's report distinguished between therapeutic and reproductive cloning and outlined key concerns by trying to respond to many as yet unresolved questions about the ethics, morality, and societal consequences of human cloning.

The council determined that the key moral and ethical objections to therapeutic cloningcloning for biological researchcenter on the moral status of developing human life. Therapeutic cloning involves the deliberate production, use, and, ultimately, destruction of cloned human embryos. One objection to therapeutic cloning is that cloned embryos produced for research are no different from those that could be used in attempts to create cloned children. Another argument that has been made is that the ends do not justify the meansthat research on any human embryo is morally unacceptable, even if this research promises cures for many dreaded diseases. Finally, there are concerns that acceptance of therapeutic cloning will lead society down the "slippery slope" to reproductive cloning, a prospect that is almost universally viewed as unethical and morally unacceptable.

The unacceptability of human reproductive cloning stems from the fact that it challenges the basic nature of human procreation, redefining having children as a form of manufacturing. Human embryos and children may then be viewed as products and commodities rather than as sacred and unique human beings. Furthermore, reproductive cloning might substantially change fundamental issues of human identity and individuality, and allowing parents unprecedented genetic control of their offspring may significantly alter family relationships across generations.

The council concluded that "the right to decide" whether to have a child does not include the right to have a child by any means possible, nor does it include the right to decide the kind of child one is going to have. A societal commitment to freedom does not require use or acceptance of every technological innovation available.

Legislation Aims to Completely Ban Human Cloning

On February 27, 2003, the U.S. House of Representatives voted to outlaw all forms of human cloning. The legislation prohibits the creation of cloned human embryos for medical research as well as the creation of cloned babies. It contains strong sanctions, imposing a maximum penalty of $1 million in civil fines and as many as ten years in jail for violations. The measure did not pass in the Senate, which was closely divided about whether therapeutic cloning should be prohibited along with reproductive cloning. In early February 2003 President Bush issued a policy statement that strongly supported a total ban on cloning. In the Senate two bills were introduced: S. 245 was a complete ban intended to amend the Public Health Service Act to prohibit all human cloning, and S. 303 was a less sweeping measure that also prohibited cloning but protected stem cell research. S. 245 was referred to the Senate Committee on Health, Education, Labor and Pensions and S. 303 was referred to the Senate Committee on the Judiciary. Neither bill, nor any comparable proposed legislation, has emerged from the Senate committees.

Even though nearly all lawmakers concur that Congress should ban reproductive cloning, many disagree about whether legislation should also ban the creation of cloned human embryos that serve as sources of embryonic stem cells. Many legislators agree with scientists that stem cells derived from cloned human embryos have medical and therapeutic advantages over those derived from conventional embryos or adults. Those who oppose the legislation calling for a total ban assert that the aim of allowing research is to relieve the suffering of people with degenerative diseases. They say that the bill's sponsors are effectively thwarting advances in medical treatment and biomedical innovation.

Supporters of the total ban contend that Congress must send an unambiguous message that cloning research and experimentation will not be tolerated. They consider cloning immoral and unethical, fear unintended consequences of cloning, and feel they speak for the public when they assert that it is not justifiable to create human embryos simply for the purpose of experimenting on them and then destroying them.

The fact that there are only about twenty available stem cell lines prompted the introduction of bills during the spring of 2004 that would require funding for human embryonic stem cell research, despite the president's 2001 policy. On April 28, 2004, more than 200 members of the House sent a letter to the president arguing in favor of an expansion of existing policy. Fifty-eight senators sent a similar letter on June 4, 2004. Pleas from patient advocacy groupsalong with the death of the former president Ronald Reagan from Alzheimer's disease on June 5, 2004, and Nancy Reagan's appeals to expand the policyfocused considerable media attention on the issue during the summer of 2004, but no legislation was passed that year.

On May 24, 2005, the House passed H.R. 810, the Stem Cell Research Enhancement Act of 2005, which would have permitted federal funding for embryonic stem cell research on cells "derived from human embryos that have been donated from in vitro fertilization clinics, were created for the purposes of fertility treatment, and were in excess of the clinical need of the individuals seeking such treatment." The Senate passed the bill on July 18, 2006, and the following day President Bush vetoed the bill.

State human cloning laws, April 2006
State Statute citation Summary Prohibits reproductive cloning Prohibits therapeutic cloning Expiration
Arizona HB 2221 (2005) Bans the use of public monies for reproductive or therapeutic cloning Prohibits use of public monies Prohibits use of public monies
Arkansas §20-16-1001 to 1004 Prohibits therapeutic and reproductive cloning; may not ship, transfer or receive the product of human cloning; human cloning is punishable as a class C felony and by a fine of not less than $250,000 or twice the amount of pecuniary gain that is received by the person or entity, which ever is greater yes yes
California Business And Professions §16004-5 Health & Safety §24185, §24187, §24189, §12115-7 Prohibits reproductive cloning; permits cloning for research; provides for the revocation of licenses issued to businesses for violations relating to human cloning; prohibits the purchase or sale of ovum, zygote, embryo, or fetus for the purpose of cloning human beings; establishes civil penalties yes no
Connecticut 2005 SB 934 Prohibits reproductive cloning, permits cloning for research; punishable by not more than one hundred thousand dollars or imprisonment for not more than ten years, or both yes no
Indiana 2005 Senate Enrolled Act No. 268 Prohibits reproductive and therapeutic cloning; allows for the revocation of a hospital's license involved in cloning; specifies that public funds may not be used for cloning; prohibits the sale of a human ovum, zygote, embryo or fetus yes yes
Iowa 707B.1 to 4 Prohibits human cloning for any purpose; prohibits transfer or receipt of a cloned human embryo for any purpose, or of any oocyte, human embryo, fetus, or human somatic cell, for the purpose of human cloning; human cloning punishable as class C felony; shipping or receiving punishable as aggravated misdemeanor; if violation of the law results in pecuniary gain, then the individual is liable for twice the amount of gross gain; a violation is grounds for revoking licensure or denying or revoking certification for a trade or occupation yes yes
Maryland 2006 SB 144 Prohibits reproductive cloning; prohibits donation of oocytes for state-funded stem cell research but specifies that the law should not be construed to prohibit therapeutic cloning; prohibits purchase, sale, transfer or obtaining unused material created for in vitro fertilization that is donated to research; prohibits giving valuable consideration to another person to encourage the creation of in vitro fertilization materials solely for the purpose of research; punishable by up to three years in prison; a maximum fine of $50,000 or both yes no
Massachusetts 2005 SB 2039 Prohibits reproductive cloning; permits cloning for research; prohibits a person from purchasing, selling, transferring, or obtaining a human embryonic, gametic or cadaveric tissue for reproductive cloning; punishable by imprisonment in jail or correctional facility for not less than five years or more than ten years or by or by imprisonment in state prison for not more than ten years or by a fine of up to one million dollars; in addition a person who performs reproductive cloning and derives financial profit may be ordered to pay profits to commonwealth yes no
Michigan §§333.2687-2688, §§333.16274-16275, 333.20197, 333.26401-26403, 750.430a Prohibits human cloning for any purpose and prohibits the use of state funds for human cloning; establishes civil and criminal penalties yes yes
Missouri §1.217 Bans use of state funds for human cloning research which seeks to develop embryos into newborn child Prohibits the use of state funds no
New Jersey §2C:11A-1, §26:2Z-2 Permits cloning for research; prohibits reproductive cloning, which is punishable as a crime in the first degree; prohibits sale purchase, but not donation, or embryonic or fetal tissue, which is punishable as a crime in the third degree and a fine of up to $50,000 yes no

State Human Cloning Laws

As of 2006 fifteen states had enacted legislation that addresses human cloning. (See Table 8.1.) California was the first state to ban reproductive cloning in 1997. Since then, twelve other statesArkansas, Connecticut, Indiana, Iowa, Maryland, Massachusetts, Michigan, Rhode Island, New Jersey, North Dakota, South Dakota, and Virginiahave passed laws prohibiting reproductive cloning. Arizona's and Missouri's legislation addresses the use of public funds for cloning, and Maryland's prohibits the use of state stem cell research funds for reproductive cloning and possibly therapeutic cloning, depending the interpretation of the statute. Louisiana also enacted legislation that prohibited reproductive cloning, but the law expired in July 2003. The laws of Arkansas, Indiana, Iowa, Michigan, North Dakota, and South Dakota also prohibit therapeutic cloning. Virginia's legislation may be interpreted as a complete ban on human cloning; however, it is unclear because the law does not define the term human being, which is used in the definition of human cloning. Rhode Island's law does not prohibit cloning for research, and California's and New Jersey's laws specifically permit cloning for the purpose of research.

State human cloning laws, April 2006 [continued]
State Statute citation Summary Prohibits reproductive cloning Prohibits therapeutic cloning Expiration
Source: "State Human Cloning Laws," National Conference of State Legislatures, April 18, 2006, (accessed October 30, 2006)
North Dakota §12.1-39 Prohibits reproductive and therapeutic cloning; transfer or receipt of the product of human cloning; transfer or receipt, in whole or in part, any oocyte, human embryo, human fetus, or human somatic cell, for the purpose of human cloning; cloning or attempt to clone punishable as a class C felony; shipping or receiving violations punishable as class A misdemeanor yes yes
Rhode Island §23-16.4-1 to 4-4 Prohibits human cloning for the purpose of initiating a pregnancy; for a corporation, firm, clinic, hospital, laboratory, or research facility, punishable by a civil penalty punishable by fine of not more than $1,000,000, or in the event of pecuniary gain, twice the amount of gross gain, whichever is greater; for an individual or an employee of the firm, clinic, hospital, laboratory, or research facility acting without the authorization of the firm, clinic, hospital, or research facility, punishable by a civil penalty punishable by fine of not more than $250,000, or in the event of pecuniary gain, twice the amount of gross gain, whichever is greater yes no July 7, A2010
South Dakota §34-14-27 Prohibits reproductive and therapeutic cloning; transfer or receipt of the product of human cloning; transfer or receipt, in whole or in part, any oocyte human embryo, human fetus, or human somatic cell, for the purpose of human cloning; cloning or attempt to clone is punishable as a felony and a civil penalty of gross gain, or any intermediate yes yes
Virginia §32.1-162.32-2 Prohibits reproductive cloning; may prohibit therapeutic cloning but it is unclear because human being is not defined in the definition of human cloning; human cloning defined as the creation of or attempt to create a human being by transferring the nucleus from a human cell from whatever source into an oocyte from which the nucleus has been removed; also prohibits the implantation or attempted implantation of the product of somatic cell nuclear transfer into an uterine environment so as to initiate a pregnancy; the possession of the product of human cloning; and the shipping or receiving of the product of a somatic cell nuclear transfer in commerce for the purpose of implantation of such product into an uterine environment so as to initiate a pregnancy. The law establishes civil penalty not to exceed $50,000 for each incident. yes unclear

California Leads the Way

In 2002 the California state legislature passed a law encouraging therapeutic cloning. Even though there were no provisions for funds in the law, the move was interpreted as support for the research. In 2004 stem cell research advocates offered voters a sweeping ballot measureProposition 71to make public funding available to support stem cell research and therapeutic cloning. Proposition 71 was championed by Robert Klein, a wealthy real estate developer and father of a child with diabetes who might benefit from the research. It also received considerable financial support from the Microsoft founder Bill Gates to finance campaign advertising and lobbying.

On November 2, 2004, Californians approved Proposition 71, a ballot measure with the potential to make the state a leader in human embryonic stem cell research. Proposition 71 enabled the state to establish its own research institutethe California Institute for Regenerative Medicine. The proposition prohibits reproductive cloning but funds human cloning projects designed to create stem cells and allocates $3 billion over ten years in research funds. Those supporting the legislation hoped that stem cell research would become the biggest, most important, and most profitable medical advancement of the twenty-first century. The legislation's supporters intended to use the funds to attract top researchers to the state, making California the epicenter of groundbreaking, lifesaving, and potentially lucrative medical research.

Nicholas Wade reports in "Plans Unveiled for State-Financed Stem Cell Work in California" (New York Times, October 5, 2006) that in October 2006 the California Institute for Regenerative Medicine released its ten-year plan for spending the $3 billion allocated to it. The institute said it will spend $823 million on basic stem cell research, $899 million on applied or preclinical research, and $656 million to advance new treatments through clinical trials. An additional $273 million will enable universities to construct laboratories in which none of the equipment has been purchased with federal funds to ensure that the researchers are not violating the rules that restrict federal money to conduct stem cell research.

Public Opinions about Stem Cell Research and Cloning

According to Gallup poll data, more than 60% of Americans believe using stem cells derived from human embryos in medical research is morally acceptable. Figure 8.5 reveals that the percentage of Americans that considers stem cell research morally acceptable had increased from 52% in 2002 to 61% in 2006.

The percentage of Americans that deems stem cell research morally acceptable varies by political affiliation, with support highest among Democrats (68%) and Independents (62%), compared with Republicans (51%). (See Figure 8.6.) According to Lydia Saad in Stem Cell Veto Contrary to Public Opinion (Gallup Poll, July 20, 2006), support also varies by educational attainmentthree-quarters (77%) of those with postgraduate degrees consider this research acceptable, compared with 45% of people who had attained a high school education or less.

The Gallup poll also found that most Americans (58%) disapproved of President Bush's July 2006 veto of a bill that would have expanded federal funding for embryonic stem cell research. (See Figure 8.7.) However, Saad notes that just 11% of Americans favor unrestricted government funding of embryonic stem cell research and another 42% support easing current restrictions. Nearly one-quarter (24%) approve of the current funding restrictions and 19% oppose any government funding of this research.

Even though Americans continue to feel that it is morally unacceptable to clone humans, public support for cloning animals increased slightly from 31% in 2001 to 35% in 2005. (See Figure 8.8.) Furthermore, unlike stem cell research, which is favored by more Democrats than Republicans; more Republicans (31%) than Democrats (28%) consider cloning animals morally acceptable. (See Figure 8.9 and Figure 8.10.)

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Cloning Genes

Cloning Genes

Gene cloning, or molecular cloning, has several different meanings to a molecular biologist. A clone is an exact copy, or replica, of something. In the literal sense, cloning a gene means to make many exact copies of a segment of a DNA molecule that encodes a gene. This is in marked contrast to cloning an entire organismregenerating a genetically identical copy of the organismwhich is technically much more difficult (with animals) and can involve ethical ramifications not associated with gene cloning. Molecular biologists exploit the replicative ability of cultured cells to clone genes.

Purposes of Gene Cloning

To study genes in the laboratory, it is necessary to have many copies on hand to use as samples for different experiments. Such experiments include Southern or Northern blots, in which genes labeled with radioactive or fluorescent chemicals are used as probes for detecting specific genes that may be present in complex mixtures of DNA.

Cloned genes also make it easier to study the proteins they encode. Because the genetic code of bacteria is identical to that of eukaryotes , a cloned animal or plant gene that has been introduced into a bacterium can often direct the bacterium to produce its protein product, which can then be purified and used for biochemical experimentation. Cloned genes can also be used for DNA sequencing, which is the determination of the precise order of all the base pairs in the gene. All of these applications require many copies of the DNA molecule that is being studied.

Gene cloning also enables scientists to manipulate and study genes in isolation from the organism they came from. This allows researchers to conduct many experiments that would be impossible without cloned genes. For research on humans, this is clearly a major advantage, as direct experimentation on humans has many technical, financial, and ethical limitations.

Cloning Techniques

Cloning genes is now a technically straightforward process. Usually, cloning uses recombinant DNA techniques, which were developed in the early 1970s by Paul Berg, of Stanford University, and, independently, by Stanley Cohen and Herbert Boyer, of Stanford and the University of California. These researchers devised methods for excising genes from DNA at precise positions, using restriction enzymes and then using the enzyme known as DNA ligase to splice the resulting gene-containing fragment into a plasmid vector .

Plasmids are small, circular DNA molecules that occur naturally in many species of bacteria. The plasmids naturally replicate and are passed on to future generations of bacterial cells. To replicate, all plasmids must contain a sequence, called an origin of replication, which directs the bacterial DNA polymerase to replicate the DNA molecule. In addition, recombinant plasmids contain one or more selectable markers. A selectable marker is a gene that confers on the bacterium harboring the plasmid the ability to survive under conditions in which bacteria lacking the plasmid would otherwise die. Usually, such genes encode enzymes that enable the bacterium to live and grow despite the presence of an antibiotic drug.

The recombinant plasmid is then introduced into a host cell, such as an Escherichia coli bacterium, by a process called transformation, and the cell is allowed to multiply and form a large population of cells. Each of these cells harbors many identical copies of the recombinant plasmid. The cells are then cultured in growth media containing the antibiotic to which the plasmid confers resistance. This ensures that only cells containing the recombinant plasmid will survive and replicate. A researcher then harvests the cells and can extract and purify many copies of the plasmid.

Another method to produce many copies of a DNA molecule, which is even simpler than traditional recombinant cloning methods, is the polymerase chain reaction (PCR). PCR amplifies the DNA in a reaction tube without the need for a plasmid to be grown in bacteria.

Importance for Medicine and Industry

The ability to clone a gene is not only valuable for conducting biological research. Many important pharmaceutical drugs and industrial enzymes are produced from cloned genes. For example, insulin, clotting factors, human growth hormone, cytokines (cell growth stimulants), and several anticancer drugs in use are produced from cloned genes.

Before the advent of gene cloning, these proteins had to be purified from their natural tissue sources, a difficult, expensive, and inefficient process. Using recombinant methods, biomedical companies can prepare these important proteins more easily and inexpensively than they previously could. In addition, in many cases the product that is produced is more effective and more highly purified. For example, before the hormone insulin, which many diabetes patients must inject, became available as a recombinant human protein, it was purified from pig and cow pancreases. However, pig and cow insulin has a slightly different amino acid sequence than the human hormone. This sometimes led to immune reactions in patients. The recombinant human version of the hormone is identical to the natural human version, so it causes no immune reaction.

Gene cloning is also used to produce many of the molecular tools used to study genes. Even restriction enzymes, DNA ligase, DNA polymerases, and many of the other enzymes used for recombinant DNA methods are themselves, in most cases, produced from cloned genes, as are enzymes used in many other industrial processes.

Genomic Versus cDNA Clones

A gene can take varying forms, and so can gene clones. The proteincoding regions of most eukaryotic genes are interrupted by noncoding sequences called introns, which are ultimately excluded from the mature messenger RNA (mRNA) after the gene is transcribed. In addition to the protein-coding sequences, all genes contain "upstream" and "downstream" regulatory sequences that control when, in which tissues, and under what circumstances the gene is transcribed. A clone containing the entire region of a gene as it exists on the chromosome, including introns and nontranscribed regulatory sequences, is called a genomic clone because it is derived directly from genomic, or chromosomal, DNA.

It is also possible to clone a gene directly from its messenger RNA transcript, from which all introns have been removed. This type of clone, called a complementary DNA or cDNA clone, includes only the protein-coding sequences and upstream and downstream sequences that do not code for amino acids but that may control how the mRNA transcript gets translated to protein.

To prepare cDNA a researcher starts with mRNA and then makes a complementary single-stranded DNA copy using the enzyme reverse transcriptase. Reverse transcriptase is a DNA polymerase that synthesizes DNA based on an RNA template that is produced by retroviruses. After the mRNA strand is digested away by another enzyme, called RNase H, DNA polymerase can synthesize a second DNA strand by using the newly made first strand cDNA as a template.

Because cDNAs lack introns, the protein-coding region in a cDNA molecule is contained in a single uninterrupted sequence, called an open reading frame, or ORF. This makes cDNA clones extremely useful for predicting the amino acid sequence of the protein that a gene encodes. It also makes it possible to direct protein synthesis from a eukaryotic cDNA clone in a bacterium, which cannot splice introns. With introns still present in a cloned gene, the bacteria will misinterpret the intron sequences as protein-encoding sequences. The resulting incorrect messanger RNA will encode a protein with an incorrect amino acid.

"Gene Cloning" Usually Means "Gene Identification"

When researchers report in a scientific journal that they have "cloned a gene" they are not referring to the rather mundane process of amplifying copies of a DNA molecule. What they are really talking about is the molecular identification of a previously unknown gene, and determination of its precise position on a chromosome. There are many different methods that can be used to identify a gene. Two of the most common approaches are discussed below.

A gene can be defined in several ways. In fact, the concept of the gene is undergoing a re-evaluation as scientists are analyzing the complete genomes of more and more organisms and finding that many sequences encode more than one protein product. Gregor Mendel identified genesfor example, he identified the factor that made peas either yellow or greenlong before he or anyone else knew that genes were encoded on segments of the DNA that made up chromosomes. Studying genetics in the fruit fly, Drosophila melanogaster, Morgan and Sturtevant demonstrated that genes are entities that reside at measurable locations, or loci, on chromosomes, although they did not yet understand the biochemical nature of genes.

Modern geneticists often use the same methods as Mendel and Morgan to identify genes by physical traits, or phenotypes, that mutations in them can cause in an organism. But today we can go even further. Using a broad range of molecular biology techniques, including gene cloning, researchers can now determine the precise DNA coding sequence that corresponds to a particular phenotype . This capability is tremendously powerful, because discovering the gene responsible for a trait can help humankind understand the cellular and biochemical processes underlying the trait. For example, geneticists have learned a great deal about the basis of cancer by identifying genes that, when mutated, contribute to cancer. By studying these genes, researchers now know that many of them control when cells divide (e.g., proto-oncogenes and tumor suppressor genes) or when they die (e.g., the apoptosis genes). Under some circumstances, when such genes are damaged by mutation, cells divide when they shouldn't, or don't die when they should, leading to cancer.

Positional Cloning

Positional cloning starts with the classical methods developed at the turn of the twentieth century by Thomas Hunt Morgan, Alfred Sturtevant, and their colleagues, of genetically mapping a particular phenotype to a region of a chromosome. A detailed discussion of genetic mapping is beyond the scope of this section, but, in general, it is based on conducting genetic crosses between individuals with two different mutant traits and analyzing how often the traits occur together in the progeny of subsequent generations.

Genetic mapping provides a general idea of where a gene is located on a particular chromosome, but it does not identify the precise DNA sequence that encodes the gene. The next step is to locate the gene on what is called the physical map of the chromosome. A physical map is a high-resolution map of all the DNA sequences that make up a chromosome. One type of physical map is a restriction map, which depicts the order of DNA fragments produced when a large DNA molecule is cut with restriction endonucleases (restriction enzymes).

Restriction maps have been made for the complete genomes of several model genetic organisms, such as the fruit fly (Drosophila melanogaster ), and the roundworm, (Caenorhabditis elegans ). For these organisms, individual large DNA fragmentson the order of forty to one hundred thousand base pairs from the whole genomehave been cloned in bacterial plasmid vectors to make a "library" of the genome. Each fragment is mapped to a known position, but the identify of the gene or genes it contains is originally unknown. To identify the genes, a cloned fragment is introduced into a mutant fly or roundworm.

To pinpoint the location of a particular gene, a researcher can introduce one or several of the plasmid clones from the physical map that are in the general vicinity of the region on the genetic map where the gene is thought to lie into a mutant that is defective in the gene of interest. If the introduced DNA corrects the mutant's defect, that DNA probably contains a normal copy of the defective gene. But these large clones usually contain several genes. By further "trimming" the DNA into smaller subfragments and testing the ability of each subfragment to rescue mutants, the researcher can eventually home in on the gene. As further confirmation that this gene is the cause of the mutant phenotype, the researcher can isolate the corresponding gene from the mutant and determine its DNA sequence to see if it contains a mutation (a DNA sequence alteration) relative to the normal gene sequence.

Expression Cloning

In some cases, a researcher becomes interested in studying a gene not because mutations in it cause an interesting phenotype but because the protein it encodes has interesting properties. A prominent example is beta-amyloid protein, which accumulates in the brains of Alzheimer's disease patients.

Expression cloning is a method of isolating a gene by looking for the protein it encodes. If the protein of interest is an enzyme, it can be found by testing for its biochemical activity. A very common method for identifying a particular protein is by using antibodies, or immunoglobulins, that bind specifically to that protein. Expression cloning usually uses a cDNA library, in which protein-coding sequences are uninterrupted by introns. Each cDNA is inserted into an "expression vector," which contains all the necessary signals for the DNA to be transcribed into mRNA. The mRNA can then be translated into protein. Thus the host cell harboring the clone will produce the gene's protein product, and the protein can then be detected by biochemical or immunologic methods. Once the cell making the protein is found, the cDNA can be re-isolated and the gene sequenced by standard means.

Gene cloning techniques continue to advance rapidly, aided by the Human Genome Project and bioinformatics. It is likely that positional cloning will take on a secondary role, and that bioinformatics and proteomics methods will begin to contribute more, as more progress in these fields is made.

see also Bioinformatics; Blotting; Chromosomes, Artificial; Cloning Organisms; Cloning: Ethical Issues; DNA Libraries; Gene; Gene Discovery; Human Genome Project; Linkage and Recombination; Marker Systems; Morgan, Thomas Hunt; Plasmid; Polymerase Chain Reaction; Recombinant DNA; Restriction Enzymes; Reverse Transcriptase; RNA Processing; Sequencing DNA; Transformation.

Paul J. Muhlrad


Alberts, Bruce, et al. Molecular Biology of the Cell, 4th ed. New York: Garland Science, 2002.

Lodish, Harvey, et al. Molecular Cell Biology, 4th ed. New York: W. H. Freeman and Company, 2000.

Micklos, David A., and Greg A. Freyer. DNA Science: A First Course in Recombinant DNA Technology. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press, 1990.

Watson, James D., et al. Recombinant DNA, 2nd ed. New York: Scientific American Books, 1992.

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Cloning Organisms

Cloning Organisms

There are two distinct types of cloning: molecular and organismal. Molecular cloning is the removal of a stretch of DNA, usually a gene, from an organism, and its insertion into another piece of DNA, such as a plasmid , to form a substance called recombinant DNA. This recombinant DNA may then be expressed in, or simply carried passively by, another organism, such as bacteria. Organismal cloning, the subject of this entry, is the production of genetically identical organisms and, as such, can be used to produce genetically identical copies of livestock or may be used to produce new members of endangered or even extinct species. It may be especially cost-effective to clone animals that produce therapeutic proteins such as blood clotting factors, thus combining both types of cloning. Cloning is controversial, however, because our understanding of the procedures needed to clone mammals may be applied to human cloning, which gives rise to profound ethical issues.

The History of Cloning

Cloning has a long history. Animals that reproduce sexually produce clones whenever identical twins are born. These twins are genetically indistinguishable, and are formed when a fertilized egg separates at a very early stage of development. Clones are also the natural product of asexual reproduction, although in this case perfect clones cannot be maintained through an infinite number of generations, because spontaneous mutations can and do occur. Lastly, clones can be produced by regeneration in both plants and animals. For example, plant cuttings will regenerate roots and, ultimately, an entire "new" plant, and some invertebrates, such as planaria, can regenerate two identical animals if the adult is cut in half. In these forms, cloning has been with us for a very long time.

Since the mid-1960s, scientists have been able to culture plant cells, that is, grow cells from plants such as tobacco and carrots in a petri dish, to get thousands of genetically identical cells. From such cultured cells an unlimited quantity of cloned plants can then be grown. These cultured cells can be modified to contain recombinant, or cloned, DNA as well.

Cloning Amphibians

The first cloning of a vertebrate by nuclear transfer was reported by John Gurdon of the University of Cambridge in the 1950s. In nuclear transplantation, the nucleus of an unfertilized donor egg is either mechanically removed or it is destroyed by ultraviolet light in a process called enucleation. The original nucleus is then replaced by a nucleus containing a full set of genes that has been taken from a body cell of an organism. This procedure eliminates the need for the fertilization of an egg by a sperm.

The most successful nuclear transplants have been achieved after serially transferring donor intestinal nuclei, that is, putting an adult nucleus from an intestinal cell into an egg whose nucleus was destroyed, allowing the egg to divide only a certain number of times, removing nuclei from these cells, and repeating this process several times before allowing the embryo to complete development. Eventually, transplantation of nuclei from albino embryonic frog cells into enucleated eggs from a dark green female frog led to the production of adult albino frog clones, demonstrating that a properly treated adult nucleus could support the full development of an egg into an adult clone. Later experiments demonstrated that nuclei from cells of other tissues, even quiescent cells such as blood cells, could also be used if properly treated. Despite these successes, no adult frog has been cloned when a nucleus from an adult cell was used without serial transfer. Without serial transfer of the nuclei, the animals would only develop to the tadpole stage, and then they would die.

Cloning of Mammals: Dolly

Nuclear transplantation has also been successful in producing mammalian clones, most notably of sheep, cattle, pigs, and mice. The most famous cloned mammal is a sheep named "Dolly," the first animal to be cloned directly from an adult cell. Experiments leading to the birth of Dolly were done at the Roslin Institute with collaborators at Pharmaceutical Proteins Limited, both in Scotland. This group had earlier produced Megan and Morag, the first mammals to be cloned from cultured cells. These two sheep were produced from embryonic cells, however, not from cells of an adult animal.

Dolly was born in the summer of 1996, the product of a nucleus from the mammary gland of a six-year-old female Finn-Dorsett sheep and an egg from a Scottish Blackface female. Mammary gland cells were grown in a petri dish and were deprived of nutrients so that they would stop dividing, just like an unfertilized egg. Donor eggs were taken from sheep soon after ovulation , and nuclei were mechanically removed from them. These enucleated eggs were then fused with the cultured mammary gland cells so that a mammary gland nucleus would be inside an unfertilized egg. Two hundred and seventy-seven such embryos were constructed and temporarily allowed to divide in a petri dish, and then all of them were transferred into the oviduct of a temporary surrogate mother. Of the original 247 embryos, only 29 developed further, and these were transferred to 13 hormonally treated surrogate mothers.

Only one surrogate mother became pregnant, and she only had one live lamb, named Dolly. The success rate was very low, but Dolly has been proven to be a true clone: She has all the characteristics of a Finn-Dorsett sheep. Independent scientists used a technique called DNA fingerprinting to show that Dolly's DNA matched the donor mammary cells but did not match that of other sheep in the Finn-Dorsett flock, nor did her DNA match that of her surrogate mother or the egg donor. Similar results have been obtained by Ryuzo Yanagimachi at the University of Hawaii, who worked with several generations of cloned mice.

In 1997 Polly, a sheep created with a combination of both molecular and organismal cloning techniques, was born. Polly was derived from a fetal sheep cell that had been engineered to contain the human gene that makes coagulation factor IX. Factor IX is missing in people with a disease called hemophilia type B. Polly and two other sheep were engineered to produce factor IX in their milk, thus providing people with hemophilia access to a safer and less expensive source of clotting factor than was previously available. Because Polly was made from more easily cultured and, therefore, more easily engineered embryonic cells, it is thought that this type of cloning technology holds the most promise for the future of pharmaceutical production of proteins that cannot be made in bacteria.

In January 2001, the first cloned member of an endangered species was born. This was a gaur, a wild ox native to India and southeast Asia, which the researchers named Noah. The gaur was chosen by Advanced Cell Technology as a candidate for cloning after the company had successfully cloned domestic cattle, which are related to the gaur species.

The embryo from which Noah developed was created from the nuclei of frozen skin cells that had been taken from an adult male gaur that had died eight years earlier. Skin cell nuclei were fused with enucleated domestic cow eggs to produce forty embryos. One of these forty was carried to full term in a surrogate cow mother. Unfortunately, Noah died of an infection two days after his birth (the infection is thought to be unrelated to his origin as a cloned animal). Despite Noah's death, it is likely that cloning will eventually be used to aid the conservation of endangered species. In the future, scientists may attempt to clone a recently extinct species, should intact DNA for an extinct species be obtained.

Problems with Cloning

In general, the success rate of mammalian cloning is low, with less than 0.1 to 2.0 percent of transplanted nuclei yielding a live birth. The vast majority of transplants fail to divide or to develop normally, indicating there is much we still do not understand about reprogramming an adult nucleus to support embryonic development. One thing that is clear, however, is that having both the donor cell and host egg cell in a nondividing state is essential for success.

What might be both the most vexing and most interesting problem with cloning is related to aging. Chromosomes "show their age" by a shortening in their tips, or telomeres , a process that occurs every time the cell they are in divides. This telomere shortening occurs in all cells except eggs, sperm, and most cancer cells, and shortened telomeres are correlated with the aging of organisms. Since the nuclear DNA in most cloned animals is taken from an adult, the chromosomes of cloned animals are expected to have shorter telomeres than animals of the same birth age that are produced by sexual reproduction, causing researchers to wonder whether cloned animals will age prematurely. Shorter telomeres have been found in Dolly and other cloned sheep, but telomeres are reported not to be shorter in cloned mice or cattle. Underlying reasons for the different results may include differences between cell types or species used.

The Myth of the Perfect Clone

Cloned animals are not 100 percent identical to their "parents." Whenever nuclear transplantation is used to produce cloned organisms, the offspring display some differences from the organism that donated the nuclei. The egg donor contributes mitochondria, the energy producers of eukaryotic cells, and these mitochondria have their own small amount of DNA-containing genes used for energy metabolism. Since mitochondria are inherited only with egg cytoplasm, they will not match the mitochondria of the animal from which the nucleus was taken. In addition, maternally derived gene products, both mRNA (messenger RNA) and protein, which serve to begin embryonic development, will differ from that of the nuclear donor, as will the uterine environment and the external environment. Thus, for example, clones produced by nuclear transplantation will be significantly less identical than will clones produced by twinning.

see also Cloning: Ethical Issues; Cloning Genes; Conservation Biology: Genetic Approaches; Hemophilia; Mitochondrial Genome; Reproductive Technology; Telomere; Transgenic Animals; Twins.

Elizabeth A. De Stasio


Gurdon, J. B., and Alan Colman. "The Future of Cloning." Nature 402 (1999): 743.

Lanza, Robert P., Betsy L. Dresser, and Philip Damiani. "Cloning Noah's Ark." Scientific American (Nov., 2000): 84-89.

Wilmut, Ian. "Cloning for Medicine." Scientific American (Dec., 1998): 58-63.

Wilmut, Ian, Keith Campbell, and Colin Tudge. The Second Creation: Dolly and the Age of Biological Control. Cambridge, MA: Harvard University Press, 2000.

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Cloning burst upon the scene in February, 1997, with the announcement of the birth of Dolly, the cloned sheep. She was created when researchers took the DNA nucleus from a cell of an adult sheep and fused it with an egg from another sheep. Shortly after Dolly was born, mice, cattle, goats, pigs, and cats were also cloned.

For biologists, however, the word cloning refers not to producing new animals but rather to copying DNA, including short segments such as genes or parts of genes. This ability to copy DNA is a basic technique of genetic engineering used in almost every form of research and biotechnology. In Dolly, copying was taken to the ultimate scale, the copying of the entire nucleus or the entire genome of the sheep. The transfer of the nucleus is usually called somatic cell nuclear transfer (SCNT), and this is what most people have in mind when they speak of cloning.

Dolly's birth immediately raised the question of human cloning. In principle, a human baby could be made using SCNT. The technical obstacles are, however, greater than most people recognize. Experts in the field doubt that human reproductive cloning can be safely pursued, at least for several decades. In Dolly's case, it took 277 attempts to create one live and apparently healthy sheep, a risk level that is clearly unacceptable for human reproduction. More important, the state of Dolly's health is not fully known. One fear associated with cloning is that the clone, having nuclear DNA that may be many years old, will age prematurely, at least in some respects. Mammalian procreation is a profoundly complicated process, as yet little understood, with subtlety of communication between sperm, egg, and chromosomes, which allows DNA from adults to turn back its clock and become, all over again, the DNA of a newly fertilized egg, an embryo, a fetus, and so forth through a complex developmental process. Using cloning to produce a healthy human baby who will become a healthy adult is decidedly beyond the ability of science as of 2002. Expert panels of scientists all strongly condemn the use of SCNT to produce a human baby.

Therapeutic cloning

Cloning, however, may have other human applications beside reproduction, and many scientists endorse these. Usually such applications are referred to as therapeutic cloning, but it should be noted that much research must occur before any therapy can be achieved. Especially interesting is the possibility of combining nonreproductive cloning with embryonic stem cell technologies. Human embryonic stem cells, first isolated in 1998, appear promising as a source of cells that can be used to help the human body regenerate itself. Based on research performed in mice and rats, scientists are optimistic that stem cells may someday be implanted in human beings to regenerate cells or tissues, perhaps anywhere in the body, possibly to treat many conditions, ranging from diseases such as Parkinson's to tissue damage from heart attack.

Embryonic stem cells are derived from embryos, which are destroyed in the process. Some scientists are hopeful that they will be able to find stem cells in the patient's own body that they can isolate and culture, then return to the body as regenerative therapy. Others think that stem cells from embryos are the most promising for therapy. But if implanted in a patient, embryonic stem cells would probably be rejected by the patient's immune system. One way to avoid such rejection, some believe, is to use SCNT. An embryo would be created for the patient using the patient's own DNA. After a few days, the embryo would be destroyed. The stem cells taken from the embryo would be cultured and put into the patient's body, where they might take up the function of damaged cells and be integrated into the body without immune response.

Religious concerns about cloning

While many believe the potential benefits justify research in therapeutic cloning, some object on religious grounds. Many Roman Catholic and Orthodox Christians reject this whole line of research because it uses embryos as instruments of healing for another's benefit rather than respecting them as human lives in their own right. Others believe that if nonreproductive cloning is permitted, even to treat desperately ill patients, then it will become impossible to prevent reproductive cloning, and so they want to hold the line against all human uses of SCNT. A few Protestant and Jewish groups and scholars have given limited approval to nonreproductive cloning.

Outside the United States, most countries with research in this area reject reproductive cloning but permit cloning for research and therapy. In the United States, federal funding is not available as of 2002 for any research involving human embryos. Privately funded research, however, faces no legal limits, even for reproductive cloning. In 2001, one U.S. corporate laboratory, Advanced Cell Technology, published its work, largely unsuccessful, to create human cloned embryos in order to extract stem cells. Some religious leaders object to this situation in which privately funded research is left unregulated.

When it comes to reproductive cloning, religious voices are nearly all agreed in their opposition, although they may give different reasons. Aside from a few isolated individuals, no one has offered a religious argument in support of reproductive cloning. All religious voices agree with the majority of scientists in their objection to cloning based on the medical risk that it might pose for the cloned person, who, even if born healthy, may experience developmental problems, including neurological difficulties, later in life. Until it is known that these risks are not significantly higher for the clone than for someone otherwise conceived, most scientists and ethicists agree that researchers have no right to attempt cloning.

Some religious scholars and organizations oppose cloning as incompatible with social justice. As an exotic form of medicine that benefits the rich, cloning should be opposed in favor of more basic health care and universal access to it.

Others oppose reproductive cloning because it goes against the nature of sexual reproduction, which has profound benefits for a species. Human beings are sexual beings, it is argued, and the necessity of sex for procreation is grounded in hundreds of millions of years of evolution and should not be lightly cast aside by technological innovation. Transcending the biological advantage of sexual procreation, some argue, are the moral and spiritual advantages of the unity of male and female in love, from which a new life emerges from the openness of being, far more than from the designs of will.

Some believe that cloning would confuse and probably subvert relationships between parents and their cloned children. If one person in a couple were the source of the clone's DNA, at a genetic level that parent would be a twin of the clone, not a parent. Whether biological confusion would amount to psychological or moral disorder is of course debatable, but any test might result in tragic consequences. Furthermore, cloning creates a child with nuclear DNA that, in some way at least, is already known. This nuclear DNA begins a new life, not with the usual uncertainties of sexual recombination but through the controls of technology. Many have said that the power to create a clone gives parents far too much power to define their children's genetic identity. Unlike standard reproductive medicine, even if combined in the future with technologies of genetic modification, cloning allows parents to specify that their child will have exactly the nuclear DNA found in the clone's original. This is assuredly not to say that parents may thereby select or control their child's personality or abilities, because persons are more than genes. But some fear that by its nature cloning moves too far in the direction of control and away from the unpredictability of ordinary procreation, so far in fact that a normal parent-child relationship cannot emerge in its proper course. To move in that direction at all is to risk subverting the virtues of parenting, such as unqualified acceptance.

Finally, some have held that cloning will place an unacceptable burden on the cloned child to fulfill the expectations that motivated their cloning in the first place. The fact that the parents may have some prior knowledge of how the clone's nuclear DNA was lived by the clone's original will lead the clone to think that the parents want a child with just these traits. One can imagine that clones will believe they are accepted and loved because they fulfill expectations and not because of their own unique and surprising identity.

In time, reproductive cloning may be widely accepted, much as in vitro fertilization has become accepted. But within religious communities, opposition to cloning is so strong that it is hard to imagine that religious people will ever accept it as a morally appropriate means of human procreation. Nevertheless, despite the strength of the objections, many recognize that human reproductive cloning will occur in time, and when it does the religious concern will shift from preventing cloning to affirming the full human dignity of the clone.

See also Animal Rights; Biotechnology; DNA; Genetic Engineering; Reproductive Technology; Stem Cell Research


brannigan, michael c., ed. ethical issues in human cloning: cross-disciplinary perspectives. new york: seven bridges press, 2001.

bruce, donald, and bruce, ann, eds. engineering genesis: the ethics of genetic engineering in non-human species. london: earthscan, 1998.

cole-turner, ronald, ed. human cloning: religious responses. louisville, ky.: westminster john knox press, 1997.

cole-turner, ronald, ed. beyond cloning: religion and the remaking of humanity. harrisburg, pa.: trinity press international, 2001.

hanson, mark j., ed. claiming power over life: religion and biotechnology policy. washington, d.c.: georgetown university press, 2001.

kass, leon r., and wilson, james q. the ethics of human cloning. washington, d.c.: aei press, 1998.

mcgee, glenn, ed. the human cloning debate. berkeley, calif.: berkeley hills books, 2000.

nussbaum, m. c., and sunstein, c. r., eds. clones and clones: facts and fantasies about human cloning. new york: norton, 1998.

pence, gregory e. who's afraid of human cloning? lanham, md.: rowman and littlefield, 1998.

pence, gregory e., ed. flesh of my flesh: the ethics of cloning humans. lanham, md.: rowman and littlefield, 1998.

ruse, michael, and sheppard, aryne, eds. cloning: responsible science or technomadness? amherst, n.y.: prometheus, 2001.

ronald cole-turner

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Cloning: Ethical Issues

Cloning: Ethical Issues

Cloning is the creation of an individual that is a genetic replica of another individual. The process transfers a nucleus from a somatic nonreproductive cell into an "enucleated" fertilized egg, one that has had its own nucleus destroyed or removed. The genes in the transferred nucleus then direct the development of a complete organism from the altered fertilized egg. Two individuals who are clones have identical genes in their cell nuclei, but differ in characteristics that are acquired in other ways.

Cloning in Context

Cloning is a natural phenomenon in species as diverse as armadillos, poplar trees, aphids, and bacteria. Identical twins are clones. Biologists have been cloning some organisms, such as carrots, for decades. Attempts to clone animals have been far less successful. They began long before the February 1997 announcement of the birth of Dolly, a sheep cloned from a mammary gland cell nucleus of a six-year-old sheep.

Oxford University developmental biologist John Gurdon cloned frogs in the 1960s, but in a limited way. He showed that a nucleus from a tadpole's intestinal lining cell could be transferred to an enucleated fertilized egg and support development to adulthood, and that a nucleus from an adult cell could support development as far as the tadpole stage. However, he was unable to coax a nucleus from an adult amphibian's cell to support development all the way to adulthood. In the 1980s several companies tried to commercialize cloning of livestock from nuclei taken from embryos or fetuses. The efforts failed because the cloned animals were nearly always very unhealthy newborns and did not survive for long. Currently, livestock cloning is limited to research, although some companies offer tissue preservation services in anticipation of future advances in commercial livestock cloning. There is no reason to believe that human clones would fare any better in terms of health or survivability than most cloned animals do.

The Cloning Ban

Ethical concerns about whether an action is "right" or "wrong" are often clouded by subjectivity, emotion, and perspective. Cloning members of an endangered species, for example, is generally regarded as a positive application of the technology, whereas attempting to clone an extinct woolly mammoth from preserved tissue elicits more negative responses, including that this interferes with nature. A project at Texas A&M University, funded by a dog lover wishing to clone a beloved deceased pet, announced the first successful cloning of a domestic animal, a cat, in February 2002. Cloning pets when strays crowd shelters might be seen as unethical. A different set of ethical issues emerges when considering the cloning of humans, which a few scientists and physicians have proposed doing outside of the United States.

Bioethics is concerned with the rights of individuals, such as the right to privacy and the right to make informed medical decisions. It is difficult to see how these issues would apply to cloning, unless someone was forced or paid to provide material for the procedure, or if an individual was cloned and not informed of his or her origin. Ethical objections to cloning seem to focus more on the fact that this is not a normal way to have a baby. Accordingly, the U.S. House of Representatives voted overwhelmingly on July 31, 2001 to pass legislation that would outlaw human cloning for any reason. However, the broadness of this action may impede other types of medical research, thus introducing a different bioethical dilemma.

The legislation seeks to ban all human cloning, both "reproductive cloning" that would be used to create a baby, and "therapeutic cloning." In therapeutic cloning, a nucleus from a somatic cell is transferred to an enucleated donor egg, and an embryo is allowed to develop for a few days. Then, cells from a part of the embryo called the inner cell mass are used to establish cultures of embryonic stem cells that are genetically identical to the individual who donated the somatic cell nucleus.

If this person has a spinal cord injury or a neurodegenerative disease, the embryonic stem cells might specialize into needed neural tissue. To treat muscular dystrophy, the cells might be coaxed to differentiate into muscle-cell precursors. Such tailored embryonic stem cells would have many applications, and a person's immune system would not reject what is essentially its own tissue. Some people argue that therapeutic cloning violates the rights of early-stage embryos; others argue that banning this research violates the rights of people who might benefit from embryonic stem cell therapy.

According to the bill's ban on producing or selling "any embryo produced by human cloning," scientists caught in the act could expect a fine of up to $1 million or ten years in prison. Proposals to exempt therapeutic cloning were defeated. The criminalization of basic research is unprecedented: Before 2001, bans on using embryonic stem cells applied only to federally funded research, and work using a small number of previously existing stem cell lines was permitted. Since the 2001 ruling, some researchers have moved to nations that permit them to derive new embryonic stem cell lines. Stem cells that are normal parts of adult bodies are being investigated as alternative sources of replacement tissues.

Cloning Misconceptions

The premise that a clone is an exact duplicate of another individual is flawed, and so if the intent of cloning is to create such a copy, it simply will not work. For example, the tips of chromosomes, called telomeres , shorten with each cell division. A clone's telomeres are as short as those from the donor nucleus, which means that they are "older" even at the start of the clone's existence. DNA in the donor nucleus has also had time to mutate, that is to say, it has had time to undergo modification from its original sequence, thus distinguishing it genetically from other cells of the donor. A mutation that would have a negligible or delayed effect in one cell of a many-celled organism, such as a cancer-causing mutation, might be devastating if an entire organism develops under the direction of that nucleus. Finally, the clone's mitochondria , the cell organelles that house the reactions of metabolism and contain some genes, are those of the recipient cell, not the donor, because they reside in the cytoplasm of the egg. Mitochondrial genes, therefore, are different in the clone than they are in the nucleus donor. The consequences of nuclear and mitochondrial genes from different individuals present in the same cell are not known, but there may be incompatibilities.

Perhaps the most compelling reason why a clone is not really a duplicate is that the environment affects gene expression. Cloned calves have different color patterns, because when the animals were embryos, the cells that were destined to produce pigment moved in different ways in each calf. For humans, consider identical twins. Nutrition, stress, exposure to infectious diseases, and other environmental factors greatly influence our characteristics. For these reasons, cloning a deceased child, the application that most would-be cloners give for pursuing the technology, would likely lead to disappointment.

Bioethical concerns over cloning may be moot, because the procedure is extremely difficult to do. Dolly was one of 277 attempts; Cumulina, the first cloned mouse, was among 15 liveborn mice from 942 tries. Cloning so often fails, researchers think, because it is not a natural way to start the development of an animal. That is, the DNA in a somatic cell nucleus is not in the same state as the DNA in a fertilized ovum . The donor DNA in cloning does not pass through an organism's germ line, the normal developmental route to sperm or egg, where gene activities are regulated as a new organism develops.

Ethical objections to human cloning are more philosophical than they are practical. The very idea of cloning assumes that our individuality can be understood so well that we can duplicate it. If human cloning ever became a reality, that this is not true would become evident. After all, we are more than a mere collection of genes.

see also Biotechnology: Ethical Issues; Cloning Genes; Cloning Organisms; Mitochondrial Genome; Stem Cells; Telomere.

Ricki Lewis


Annas, George J. "Cloning and the U.S. Congress." The New England Journal of Medicine 346 (2002): 1599.

Holden, Constance. "Would Cloning Ban Affect Stem Cells?" Science 293 (2001): 1025.

Lewis, Ricki. "The Roots of Cloning." In Discovery: Windows on the Life Sciences. Medford, MA: Blackwell Science, 2000.

Mayor, Susan. "Ban on Human Reproductive Cloning Demanded." British Medical Journal 322 (Jun., 2001): 1566.

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Cloning: Applications to Biological Problems

Cloning: Applications to biological problems

Human proteins are often used in the medical treatment of various human diseases. The most common way to produce proteins is through human cell culture , an expensive approach that rarely results in adequate quantities of the desired protein. Larger amounts of protein can be produced using bacteria or yeast . However, proteins produced in this way lack important post-translational modification steps necessary for protein maturation and proper functioning. Additionally, there are difficulties associated with the purification processes of proteins derived from bacteria and yeast. Scientists can obtain proteins purified from blood but there is always risk of contamination . For these reasons, new ways of obtaining low-cost, high-yield, purified proteins are in demand.

One solution is to use transgenic animals that are genetically engineered to express human proteins. Gene targeting using nuclear transfer is a process that involves removing nuclei from cultured adult cells engineered to have human genes and inserting the nuclei into egg cells void of its original nucleus .

Transgenic cows, sheep, and goats can produce human proteins in their milk and these proteins undergo the appropriate post-translational modification steps necessary for therapeutic efficacy. The desired protein can be produced up to 40 grams per liter of milk at a relatively low expense. Cattle and other animals are being used experimentally to express specific genes, a process known as "pharming." Using cloned transgenic animals facilitates the large-scale introduction of foreign genes into animals. Transgenic animals are cloned using nuclear gene transfer, which reduces the amount of experimental animals used as well as allows for specification of the sex of the progeny resulting in faster generation of breeding stocks.

Medical benefits from cloned transgenic animals expressing human proteins in their milk are numerous. For example, human serum albumin is a protein used to treat patients suffering from acute burns and over 600 tons are used each year. By removing the gene that expresses bovine serum albumin, cattle clones can be made to express human serum albumin. Another example is found at one biotech company that uses goats to produce human tissue plasminogen activator, a human protein involved in blood clotting cascades. Another biotech company has a flock that produces alpha-1-antitrypsin, a drug currently in clinical trials for the use in treating patients with cystic fibrosis. Cows can also be genetically manipulated using nuclear gene transfer to produce milk that does not have lactose for lactose-intolerant people. There are also certain proteins in milk that cause immunological reactions in certain individuals that can be removed and replaced with other important proteins.

There is currently a significant shortage of organs for patients needing transplants. Long waiting lists lead to prolonged suffering and people often die before they find the necessary matches for transplantation. Transplantation technology in terms of hearts and kidneys is commonplace, but very expensive. Xenotransplantation, or the transplantation of organs from animals into humans, is being investigated, yet graft versus host rejection remains problematic. As an alternative to xenotransplantation, stem cells can be used therapeutically, such as in blood disorders where blood stem cells are used to deliver normal blood cell types. However, the availability of adequate amount of stem cells is a limiting factor for stem cell therapy.

One solution to supersede problems associated with transplantation or stem cell therapy is to use cloning technology along with factors that induce differentiation. The process is termed, "therapeutic cloning" and might be used routinely in the near future. It entails obtaining adult cells, reprogramming them to become stem cell-like using nuclear transfer, and inducing them to proliferate but not to differentiate. Then factors that induce these proliferated cells to differentiate will be used to produce specialized cell types. These now differentiated cell types or organs can then be transplanted into the same donor that supplied the original cells for nuclear transfer.

Although many applications of cloning technology remain in developmental stages, the therapeutic value has great potential. With technological advancements that allow scientists to broaden the applications of cloning becoming available almost daily, modern medicine stands to make rapid improvements in previously difficult areas.

See also DNA hybridization; Immunogenetics; Microbial genetics; Transplantation genetics and immunology

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cloning is the generation of genetically identical organisms: each group of such organisms is a clone. Ever since Aldous Huxley's Brave New World, cloning and clones have been the subject both of science fiction and of serious public concern over their possible biotechnological applications. Before taking a paranoid view, however, it is worth noting that clones occur widely and naturally. Many plant varieties are propagated as clones (for instance by grafting) and the summer aphids preying upon them are asexually produced, genetically identical individuals — clones. Identical twins are clones, and the famous Dionne quintuplets born in Canada in 1934 represent a human clone of five people.

Sexual reproduction involves a re-assortment of the genetic material from the two parents and hence the generation of new, genetically distinct individuals. In contrast to this, methods of asexual reproduction result in the production of genetically identical individuals. Bacteria, yeast, and the individual cells of multicellular organisms are able to reproduce asexually, and the products of such replication are clones. Thus, for instance, all the cells in a multicellular organism represent one clone derived from the fertilized egg. During the process of development, and indeed at later stages of life, there may be stably inherited restrictions on the use of the genetic material or new mutations which define new clonally-related groups of cells.

The cells of malignant tumours, for instance, usually carry numbers of mutations which were not originally present in the normal cells of the individual; as these cancer cells progress newer mutations may arise so that several discernibly different clones of cells may be found. One question of interest would be whether all the cells arise from one single event — is the tumour a clone? This question may be addressed in individuals where there is already more than one distinguishable clone of cells present. In women, one of the two X chromosomes will have been inactivated early in development in a random but stable manner. This results in all the tissues being a mosaic of two alternative types of cell. Tumours typically display a single type, demonstrating their clonal origin from a single precursor cell.

This illustrates another important aspect of cloning: the origin of the clone purifies it from a mixed population. For example, many cultivated plants are deliberately propagated asexually by cuttings or grafting, so that one particular variety may be maintained. In molecular biology, this property — that the isolation of a clone selects, maintains, and propagates as a single pure variant — is used directly for analysis of the genetic material itself; the DNA. Pieces of DNA are inserted into a bacterial or viral host in a form that replicates asexually. One single cell is used to start a colony — a clone — and thus large amounts of a single purified DNA fragment may be isolated.

All the cells of a multicellular organism arising from one fertilized egg are clones and, unless subsequently modified, contain the same genetic information. This was demonstrated in plants by regeneration of a whole plant from a single cell from a carrot root. In animals it was shown possible to transplant the nucleus from a gut cell of a tadpole into a fertilized egg, which had had its own nucleus destroyed, and regenerate a new tadpole which now had the genetics of the donor nucleus. Such cloning was first attempted for mammals using mice, but this did not work with any nuclei other than those from the earliest embryos. In the 1990s, however, Ian Wilmut and a team at the Roslin Research Institute in Edinburgh demonstrated a technique allowing nuclei from cells in tissue culture to be used to clone a sheep. They have now demonstrated that these tissue culture cells can be derived from an adult sheep.

The lamb (named Dolly), who was produced from a nucleus from a cell grown from the breast tissue of an adult sheep, has had major political impact as it is now clear that there is no theoretical reason why this cloning should not be possible not only with sheep but with other mammals, including humans. Cloning people is illegal in Britain, but world-wide legislation is not in place. In some quarters it is argued, however, that the technique per se might be useful to regenerate transplant tissues or organs without ever compromising the ethical, legal, and moral susceptibilities that would arise from deliberately generating whole fetuses or people.

Martin Evans

See also biotechnology; stem cells.

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A clone is a group of genetically identical cells descended from a single common ancestor. Cloning is one method for producing identical twins. After an egg is fertilized, it begins to divide repeatedly. If the egg completely separates during the two-cell stage, identical twins will result. Both individuals will have exactly the same combination of genes (genotype) and each will have the same physical characteristics (phenotype). This is an example of how exact duplicates can naturally occur through sexual reproduction.

Science has capitalized on the mechanisms of cellular reproduction to produce clones. Advances in biotechnology since the 1970s have enabled livestock breeders to clone virtually unlimited numbers of identical animals from a single embryo. This allows the precise duplication of an animal with desired characteristics.

In 1979 veterinarian Steen Willadsen developed a way to divide sheep embryos in half at the two-cell stage, making clones possible. In the next few years, several scientists made further strides in this area with both sheep and cattle embryos. A team developed a simplified method of dividing and cloning sheep embryos in 1984.

Cloning is one area of genetics that is advancing very rapidly, and it is therefore very controversial. If this technology is ever applied to humans, who will decide which genes are "desired" and should be cloned? This is only one of many important questions that have arisen as a result of genetic cloning.

Dairy Farmers Use Cloning Techniques

As an example of cloning techniques, dairy fanners trying to clone a cow with high milk-producing qualities begin by artificially inseminating a high-producing cow with the sperm from a prize bull. The resulting embryo, which contains the entire genetic instructions needed to form a complete calf, develops within its mother. After some time, the embryo divides into a mass of 32 identical cells. The embryo is then carefully removed from the mother cow and separated into 32 separate cells. Finally, after microsurgery on the cells, each new embryo is transplanted into 32 different carrier cows, where it develops fully.

After a normal pregnancy, each carrier cow gives birth to a calf that is genetically identical to the 31 other calves derived from the original 32 cell embryo. Each calf is a clone. The trait for increased milk production has been cloned so that the farmer now has 32 high milk-producing cows instead of just one. Cloning technology has enabled breeders to develop lines of cattle, sheep, and cotton plants that respectively produce more milk, wool, and cotton.

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Clone, Cloning


A clone is a group of organisms that derive from a single ancestor and are genetically identical. A clone can be a group of mammals such as sheep, or a group of cells in culture.

Cloning cells is a powerful tool in biology and medicine, since growing large quantities of identical cells allows for a large harvest of the various identical and useful components of these cells. It is possible to construct genetic components in the laboratory, place them in cells, and then have the cells grow and multiply to produce large quantities of the components.

Cloning is an essential technique in modern molecular biology; it is used widely in studying genetic effects in the drug-abuse field. Cloning much larger organisms such as cows and sheep is expected to have a major impact in that production of the best of any species can theoretically be accomplished by cloning. This is an important goal in agriculture today.

Michael J. Kuhar

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KUHAR, MICHAEL J.. "Clone, Cloning." Encyclopedia of Drugs, Alcohol, and Addictive Behavior. 2001. 29 May. 2016 <>.

KUHAR, MICHAEL J.. "Clone, Cloning." Encyclopedia of Drugs, Alcohol, and Addictive Behavior. 2001. (May 29, 2016).

KUHAR, MICHAEL J.. "Clone, Cloning." Encyclopedia of Drugs, Alcohol, and Addictive Behavior. 2001. Retrieved May 29, 2016 from

cloning vector

cloning vector See vector.

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"cloning vector." A Dictionary of Biology. 2004. 29 May. 2016 <>.

"cloning vector." A Dictionary of Biology. 2004. (May 29, 2016).

"cloning vector." A Dictionary of Biology. 2004. Retrieved May 29, 2016 from


cloning: see clone.

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"cloning." The Columbia Encyclopedia, 6th ed.. 2016. 29 May. 2016 <>.

"cloning." The Columbia Encyclopedia, 6th ed.. 2016. (May 29, 2016).

"cloning." The Columbia Encyclopedia, 6th ed.. 2016. Retrieved May 29, 2016 from

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