Science of cloning

Summary

Cloning is any type of biological reproduction that produces offspring that are genetically identical to their parents. Cloning occurs naturally since many organisms routinely reproduce through natural cloning processes. Artificial cloning technologies include molecular cloning, which reproduces large quantities of discrete segments of DNA; reproductive cloning, which uses assisted reproductive technologies to produce animals that share the same desirable genetic characteristics as another living or previously existing organism; and therapeutic cloning, which uses the same techniques as reproductive cloning but instead derives useful cell lines from cloned embryos.

Definition and Basic Principles

Cloning is a means of producing biological organisms, cells, or DNA molecules that are genetically identical to their progenitors. There are natural forms of cloning and three main types of artificial cloning: molecular, reproductive, and therapeutic cloning.

Natural mechanisms of cloning occur in organisms such as bacteria that simply split or fragment into identical copies of themselves. In other organisms, reproductive cells, or gametes, undergo a process called parthenogenesis, in which they initiate development without the benefit of fertilization. Cloning is uncommon in mammals, but rarely, early mammalian embryos undergo a form of cloning called twinning, in which the embryo splits into two embryos, which develop into genetically identical twins.

Molecular cloning, also known as recombinant DNA technology or DNA cloning, involves the transfer of an isolated fragment of DNA from an organism of interest to a host cell that replicates it. Such isolated DNA fragments are known as cloned DNA or genes.

Reproductive cloning uses assisted reproductive technologies to generate animals with the same nuclear genome as another animal. The particular procedure used during reproductive cloning is called somatic cell nuclear transfer (SCNT). Cloned embryos are gestated in the womb of a surrogate mother until they come to term. Cloned organisms are not genetically modified organisms but are simply produced through a type of assisted reproduction.

Therapeutic cloning uses the same procedures as reproductive cloning; however, instead of transferring the cloned embryo into the womb of a surrogate mother, the embryo is further manipulated in the laboratory to make cell cultures of embryonic cells for basic or clinical research.

Background and History

Sea urchins were the first animals cloned in the laboratory. In 1894, Hans Dreisch isolated sea urchin embryo cells and watched them develop into small, separate larvae. In 1902, Hans Spemann used the same procedure of embryo splitting to isolate cells from salamander embryos, which also developed into identical adult salamanders. In 1903, US Department of Agriculture employee Herbert Webber coined the word “clon” for asexually produced cells or organisms, which later evolved into “clone.” This term comes from the Greek klon, which means “trunk” or “branch.” Horticulturists have used this term for more than a century since an entirely new plant can grow from a cutting, resulting in a plant that is genetically identical to the plant from which the cutting was taken.

In 1928, Spemann cloned salamanders by transferring the nucleus, the subcellular compartment that houses the chromosomes, from one salamander embryo into the egg of another. Since Spemann's seminal experiments, scientists have adapted nuclear transfer technology to clone other organisms. In 1952, frogs were cloned, and in 1963, the Chinese embryologist Tong Dizhou cloned a carp to produce the first cloned fish. During the 1980s and 1990s, sheep, cows, and mice were cloned. However, all these animals were cloned by using nuclei from embryos. In 1996, Ian Wilmut and his team at the Roslin Institute in Edinburgh, Scotland, cloned a sheep from an adult cell, demonstrating that adult cells could serve as the source of genetic material for animal clones. This technological feat was followed by the cloning of goats, mules, gaurs (an endangered species), horses, pigs, mouflons (wild sheep), mice, rats, dogs, cats, water buffalos, camels, rabbits, deer, wolves, and African wildcats, and even embryos from nonhuman primates and humans.

How It Works

Molecular Cloning. To clone a gene, the DNA of the model organism is selectively fragmented by enzymes called restriction endonucleases (REs) and inserted into another piece of DNA called a cloning vector. Cloning vectors are either small circles of DNA called plasmids, bacterial viruses, or bacterial or yeast artificial chromosomes. They ferry the DNA fragments from the genome of the model organism into a host cell (either a bacterium or yeast). This population of host cells collectively carries the entire genome of the model organism in small fragments and is called a gene library.

To isolate a gene from a gene library requires a probe, which is a fragment of DNA or RNA of any length that has a sequence that is complementary to the sequence of the gene that is to be isolated. Probes can be made synthetically or can come from the genes of closely related organisms. By screening the gene library with the probe, the gene of interest is cloned, which simply means to isolate it from all the other sequences found in the genome of the model organism.

Alternatively, scientists can synthesize small strands of DNA called primers, whose sequences are complementary to different locations in the gene. These primers can be used to specifically amplify the gene from the library by means of a polymerase chain reaction (PCR). A polymerase chain reaction makes large quantities of the gene of interest from a very small amount of starting material, and the amplified DNA can also be cloned into a cloning vector or analyzed directly.

Reproductive Cloning. To clone an animal, mature eggs are isolated from females of the animal species that is to be cloned. The egg is enucleated by piercing it with a microscopically narrow (0.0002-inch-wide) glass tube that is used to vacuum out the egg nucleus. The enucleated egg is fused with a cell from the body of the animal to be cloned and activated with either chemicals or an electric current. This procedure is called somatic cell nuclear transplantation (SCNT).

After activation, the egg divides and grows like a newly formed embryo. However, if the animal is a mammal, the embryo can survive only for a limited period of time before it must implant into the inner layer of the mother's womb. Therefore, a surrogate female from the same species of the animal to be cloned, or a closely related species, is made pseudopregnant by feeding her hormones, and the embryo is released into her receptive womb, where it then implants. Barring any technical or biological mishap, the cloned embryo will develop, and the process will result in a live birth.

Therapeutic Cloning. To make embryonic cell cultures, cloned embryos are made by means of somatic cell nuclear transplantation. They are then either disassembled in the laboratory and used to establish embryonic cell cultures or gestated in a surrogate mother to the fetal stage, at which time the fetus is aborted, and cells from the fetus are used to establish fetal cell cultures.

By culturing specific cells from cloned embryos, scientists can make embryonic stem cell (ESC) cultures. During mammalian development, two distinct cell populations form after the first few days of embryonic development. The trophoblast, or the flattened outer layer of cells, will eventually form the placenta and its associated structures. The inner cell mass (ICM) is the round, inner clump of cells that develops to form the embryo proper and a few structures associated with the placenta. If ICM cells are isolated and cultured on feeder cells, a layer of non-dividing skin cells that secrete a cocktail of growth-promoting chemicals, the ICM cells will grow and spread over the surface of the culture dish. Such a culture is an embryonic stem cell culture, and these cells are pluripotent, which means that they can differentiate into any cell type in the adult body.

Applications and Products

Molecular Cloning. Organisms that express cloned genes make many useful pharmaceuticals, such as human insulin, growth hormone, clotting factors, fertility drugs, and vaccines. Cloned genes are also used to genetically screen individuals for genetic diseases. Pharmacologists even use cloned genes for pharmacogenetics, which screens patients for the presence of gene variants that can profoundly affect the efficacy and toxicity of particular drugs. This allows clinicians to tailor treatment to the exact genetic makeup of the patient to maximize treatment efficacy and minimize side effects. Such a strategy is called personalized medicine. Cloned genes are also used in gene therapy, which delivers cloned genes into the bodies of patients who suffer from genetic diseases in an attempt to cure them. Patients with cancer and inherited deficiencies of the immune system, blindness, and blood-based defects have been treated with gene therapy protocols.

In agriculture, the introduction of cloned genes into plants that are used as food crops has generated transgenic crops. These crops display several advantageous traits—reduced dependence on agrochemical applications (for example, Bt-corn and herbicide-resistant crops), increased nutritional value (for example, Golden Rice), increased resistance to environmental stresses, and reduced spoilage (for example, the Flavr Savr tomato).

Despite safety and ethics concerns regarding genetically modified organisms (GMOs) in agricultural products, the International Service for the Acquisition of Agri-Biotech Applications (ISAAA) reported in the early 2010s that genetically modified (GM) crops were planted in twenty-seven countries by approximately 18 million farmers. Over 60 percent of the world's population lived in the twenty-seven countries that were planting GM crops. In 2006, US government statistics showed that 87 percent of the global genetically modified crops were grown in developed countries. By 2013, however, ISAAA reported that Latin American, Asian, and African farmers grew 54 percent of the global GM crops compared to 46 percent in developed countries. This trend continued into 2016 when the ISAAA reported increased soybean plantings in Argentina and Brazil. Corn, soybeans, cotton, alfalfa, and canola were the major crops, often modified for insect resistance. Rice has been genetically enhanced to produce more iron and vitamins to alleviate malnutrition. Other plants have been modified to survive weather variances.

By the 2020s, over seventy countries had adopted GM crops, with around thirty producing crops and the others importing them. The ISAAA reported GM crops had increased global food, feed, and fiber production by one billion tons between 1996 and 2020. This production also reduced the environmental impact of farming by 17 percent.

Reproductive Cloning. When farmers identify food animals with desirable traits, they typically breed those animals as much as possible to improve the genetic quality of their herds and flocks. However, such prize animals inevitably die. Propagating these animals by reproductive cloning and mating them to as many animals as possible preserves the exceptional genetic content of a prize animal and allows it to produce far more offspring. This significantly raises the genetic quality of the flock or herd, and commercial dissemination of such cloned animals to other farmers raises the overall genetic quality of food animals. Reproductive cloning also eliminates the need for artificial insemination, which is often expensive and inconvenient.

Cloning effectively maintains high-quality animal stocks. Reproductive cloning of only the healthiest and most productive animals increases their numbers and improves the gene pool (total of genetic diversity) and overall health of food animals. This results in safer and healthier food and reduces the use of growth hormones, antibiotics, and other chemicals in raising animals.

In the field of conservation biology, the numbers of endangered species are often increased by captive breeding programs. However, not all endangered species can effectively breed in captivity. Reproductive cloning can aid in the preservation of those organisms that do not reproduce in captivity. Cloning can also resurrect genetic material from dead animals and potentially expand the gene pool of endangered species. In 2001, scientists at the University of Teramo, Italy, cloned the European mouflon, an endangered sheep, from cells sampled from a dead animal. When combined with other reproductive technologies, cloning can help save endangered species. Similarly, in 2023, scientists in China cloned three cows capable of producing nearly twice the amount of milk. Scientists have also successfully cloned horses, cats, dogs, ferrets, primates, frogs, and camels.

Cloned animals also serve as excellent research models. Because each cloned animal is genetically identical, experiments on cloned animals are devoid of differences caused by heterogeneous genetic backgrounds. Genetic manipulation of cloned animals allows researchers to modify genes of interest and more completely analyze their contribution to development and disease. Modifying particular genes of cloned animals also generates model systems for particular genetic diseases. Cloned, transgenic mice and cloned knockout mice, which have had a specific gene inactivated, are examples of the vast usefulness of such model systems.

Of enormous interest is modifying the genomes of cloned animals so that they can produce clinically and pharmaceutically significant products. By genetically modifying pigs, it is possible to make cloned pigs that contain organs that are fit for transplantation into humans (xenotransplantation). Also, producing antibodies, clotting factors, or even vaccines in the blood or milk of farm animals provides a means to mass-produce potentially expensive pharmaceutical agents at a fraction of the normal cost. This process is called pharming.

Therapeutic Cloning. Therapeutic cloning has tremendous potential for numerous clinical applications. Embryonic stem cells (ESCs) made from therapeutic cloning procedures are pluripotent. Therefore, injured, diseased, or failing tissues or organs could be replaced by tissues or organs manufactured from embryonic stem cells in the laboratory or fetal cells from cloned fetuses. Furthermore, embryonic stem cells made from cloned embryos, or any tissues or organs fashioned from these cells, would not be regarded by the patient's body as foreign. Experiments in laboratory animals have shown that such scenarios are possible. Therapeutic cloning, coupled with embryonic stem cell technology, could christen a new era of regenerative medicine.

Embryonic stem cells from cloned embryos have toxicological applications. Toxicologists typically use laboratory animals or cultured cells to gauge the biological effects of natural or industrially produced molecules on humans. However, laboratory animals show limited utility as a model for human toxicology, and cultured cells do not represent the response of an organ or tissue to foreign molecules. Furthermore, neither of these model systems can assess the individual responses people will have to such molecules because the genetic variation between individual humans causes differential responses to drugs, toxins, or environmental pollutants. However, cultured embryonic stem cells from cloned embryos can test the biological effects of drugs or environmental pollutants on cells made from a specific person. In addition, because these cells can be differentiated into various tissues and even organs, they can be used to evaluate the individual and tissue-specific responses people might have to particular drugs or pollutants.

Careers and Course Work

Students who wish to pursue a career in cloning should possess a foundational grasp of biology and chemistry. Advanced coursework in cell, molecular, and developmental biology is of cardinal importance. Many entry-level jobs exist in academic and industrial laboratories for those with a bachelor's degree in biology, biochemistry, or chemistry. Such jobs are usually laboratory technician positions, and good laboratory skills are required. For those who wish to work as a research group leader, a PhD in either cell or developmental biology, a DVM, or an MD degree is required.

Cloning work requires highly skilled technicians who are very dexterous and can look through microscopes for a long period while performing extremely fine manipulations. Such techniques often require many weeks of practice to perfect, and a patient, forbearing personality greatly helps people in the cloning field. Because cloning experiments are also very labor intensive, a collaborative mindset is also helpful.

Many scientists are involved in cloning work in industry, and several biotechnology companies have divisions that investigate cloning technology. Because such companies normally attempt to clone organisms for profit, cloning divisions of biotechnology companies usually examine improving the efficiency and cost-effectiveness of cloning procedures to standardize the manufacture of clones.

Many cloning scientists work in academic laboratories that devote their time to more basic research questions. Scientists in academic institutions must split their time between teaching and research. Other scientists who work for government laboratories, such as the NIH, can work on cloning, safety testing of cloned products, and other aspects of cloning without teaching responsibilities.

Social Context and Future Prospects

Though controversial, cloning plays an important role in modern science. Many foods contain genetically engineered products. Physicians prescribe medicines, give vaccines, and apply other biological products made by genetically engineered microorganisms on a quotidian basis. Genetically modified organisms have become the focal point of concern for many environmental activism groups. Such groups oppose GMOs because they believe that cloned genes can spread to other species and cause severe environmental disruption and that genetically engineered foods have not been sufficiently tested and are potentially dangerous to human health.

Though cloning can help maintain and repopulate species on the brink of extinction, this concept highlights the superficial nature of this type of conservation remedy. Conservation biologists posit that cloning endangered species does not address the habitat destruction and environmental degradation that pushed these species to extinction in the first place. Second, cloning makes one species and does not recreate an ecosystem. For example, cloning cannot recapitulate a coral reef or an old-growth forest. Thus, many argue that it is the wrong solution to the problem.

The most contentious aspects of cloning technologies are human genetic engineering and reproductive cloning. Until the cloning of the sheep Dolly in 1997, it was thought that adult specialized cells could not be made to revert to nonspecialized cells that could give rise to any type of cell. However, Dolly was created from a specialized adult cell from a ewe’s udder. US President Bill Clinton asked the National Bioethics Advisory Commission to form recommendations about the ethical, religious, and legal implications of human cloning. In June 1997, the commission concluded that attempts to clone humans were “morally unacceptable” for safety and ethical reasons, and restrictions were placed on the use of federal funds for human cloning. In January 1998, the US Food and Drug Administration (FDA) declared it had the authority to regulate human cloning and that any human cloning must have FDA approval.

In 2013, the science of cloning experienced another breakthrough when researchers at the Oregon Health and Science University successfully cloned human embryonic stem cells. Using nuclear transfer, the scientists implanted the skin of a fetus into the nucleus of an egg before prompting division, creating a ball of cells that includes embryonic stem cells with the same genetic material as the original skin cell. The primary purpose of the research was not to further the prospect of human cloning, however, but to establish a process to consistently clone stem cells for use in the treatment of diseases and other human ailments.

Transhumanists are some of the most energetic proponents of human cloning and genetic enhancement. As a movement, transhumanism regards infirmity, disease, aging, and death as undesirable and unnecessary and views science and technology as the means to defeat human limitations. Transhumanists' main argument for human cloning is that reproductive freedoms extend to everyone, and therefore, every human has an inherent right to clone themselves.

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