The History of Genetic Engineering

Genetic engineering owes its existence to the developments in molecular genetics, virology, and cytology that culminated in the determination of the structure of DNA by James Watson and Francis Crick in 1953. Building on research involving bacteriophages (a bacterial virus), Joshua Lederberg, a geneticist at the University of Wisconsin, found that bacteria can transfer genetic information through plasmids, small mobile pieces of DNA that exist independent of the chromosomes. In the 1950s, Lederberg pioneered the earliest techniques in genetic engineering, shuffling genetic material between bacterial cells. After the identification of restriction enzymes capable of "cutting" DNA in specific locations in 1968, scientists were able to insert foreign DNA directly into bacterial cells. The discovery that the foreign DNA would naturally bond with the host DNA, made it possible to splice together genes from multiple organisms, the technique used in recombinant DNA engineering. Although highly complicated, rDNA engineering can be simply explained: genetic material from the donor source is isolated and "cut" using a restriction enzyme and then recombined or "pasted" into the genetic material of the receiver. By 1971, advanced transplantation techniques had been developed and rDNA techniques using the restriction enzyme EcoRi were operable the following year, leading to the first experiments in genetic engineering.
In 1973, Stanford biochemist Stanley Cohen under-took one of the first rDNA experiments, inserting a piece of bacterial DNA into Escherichia coli (E. coli), a bacterium found in the human intestine. However, the research soon became controversial, particularly when American molecular biologist Paul Berg designed an experiment to insert DNA from simian virus #40 (sv40)—a known cancer-causing agent—into E. coli. As word of the daring procedure spread, the public was captivated and fearful, afraid that a genetically engineered virus, inured to antibiotics and carried in a common bacterium, could escape and cause an epidemic. Hoping to diffuse fears of a potential biohazard and maintain control of their research, over one hundred and fifty molecular biologists and related specialists met at the Asilomar Conference Center in Monterey, California, in late February 1975. The conference represented an extraordinary moment in the history of science, as the research community, recognizing its social responsibility, officially adopted a moratorium until appropriately safe procedures and guidelines could be developed. The conference ultimately resulted in the "National Institutes of Health Guidelines for Research Involving rDNA Molecules" and an ongoing National Institute of Health rDNA Advisory Committee (RAC)founded in 1974.

Yet the guidelines only increased public concern over genetic engineering. Critics charged that attempts to splice genes together from different organisms were akin to "playing God" and could result in dangerous and immoral hybrids. Adopting the literary example of "Dr. Frankenstein's monster" as an appropriate symbol of misguided science, opponents of rDNA engineering converged on research laboratories and public meetings. An attempt to build a recombinant laboratory at Harvard University set off such a firestorm that local politicians created a review board to assess potential risks, eventually requiring more stringent controls than those set by the NIH. By 1977, protests of rDNA facilities had spread to other campuses—the University of California San Diego, the University of Wisconsin, the University of Michigan, and the University of Indiana—while the state legislatures of New York, New Jersey, and California held public hearings. However, it was the resolution of an old court case and the introduction of a new form of rDNA engineering that ultimately created the greatest controversy.

In a monumental decision handed down on 16 June 1980, the United States Supreme Court held in Diamond v. Chakrabarty that man-made life forms were subject to patent laws and protection. The decision resolved a longstanding issue on patents and organic material, as the case dated to 1972, when Ananda Chakrabarty, a researcher at General Electric, applied for a patent on a form of Pseudomonas bacteria bred (but not genetically engineered)to digest oil slicks. By a narrow five to four margin the court construed the Patent Act, originally drafted by Thomas Jefferson, so as to include all products of human invention, relying on a 1952 Senate report that recognized as patentable "anything under the sun that is made by man." More than any other single event, the ruling galvanized many mainstream religious communities and environmental groups, eventually resulting in a letter of protest to President Carter and an indepth review by the President's Commission for the Study of Ethical Problems in Medicine and Biomedical and Behavioral Research (1980–1983). The commission's report, issued in 1982 and entitled Splicing Life: The Social and Ethical Issues of Genetic Engineering with Human Beings, emphasized the importance of rDNA engineering to biomedical progress and American industries, arguing that it was best that the research be conducted under the auspices of government regulation and control. However, while the study resolved anxiety over rDNA engineering and patenting, proponents of genetic engineering still had to address concerns over the development of "germ-line" engineering, a controversial procedure that allowed scientists to literally create new strains of organisms.

Germ-line engineering differs from rDNA engineering in that the donor genes are inserted into a "germ," or reproductive cell, thereby permanently altering the genetic makeup of the organism's descendants. For example, in 1982, Ralph Brinster of the University of Pennsylvania Veterinary School inserted the gene that produces rat growth hormone into mouse embryos. The resulting strain of mice, dubbed "super mice" by the press, expressed the gene and thus grew into a substantially larger and more powerful new breed of mouse. Critics of germ-line engineering quickly denounced the technique as immoral and argued it was a form of "anthropomorphic Lamarckism."
Jean-Baptiste de Lamarck, a nineteenth-century French naturalist, had proposed that traits acquired during an organism's lifetime were passed on to its progeny—an idea refuted by Darwinian evolutionary theory. Yet, in germ-line engineering, traits acquired during the organism's lifetime are passed on, but only those traits deemed necessary or desirous by man. Environmental groups also denounced germ-line engineering because of "biosafety" concerns, fearing that genetically engineered species, which would possess a distinct advantage over nonengineered species, could upset the globe's finely tuned ecological systems. However, because most politicians, scientists, and manufacturers believed the potential benefits from rDNA and germ-line engineering outweighed its potential dangers, the protests were overshadowed by the development of a biotechnology industry based on genetic engineering.

Animal cells and plant cells

Animal cells

In contrast to the wide variety of plasmid and phage vectors available for cloning in prokaryotic cells, relatively few vectors are available for introducing foreign genes into animal cells. The most commonly used are derived from simian virus 40 (SV40). Normal SV40 cannot be used as a vector, since there is a physical limit to the amount of DNA that can be packaged into the virus capsid, and the addition of foreign DNA would generate a DNA molecule too large to be packaged. However, SV40 mutants lacking portions of the genome can be propagated in mixed infections in which a “helper” virus supplies the missing function.

Plant cells

Two systems for the delivery and integration of foreign genes into the plant genome are the Ti plasmid of the soil bacterium Agrobacterium and the DNA plant virion cauliflower mosaic virus. The Ti plasmid is a natural gene transfer vector carried by A. tumefaciens, a pathogenic bacterium that causes crown gall tumor formation in dicotyledonous plants. A T-DNA segment present in the Ti plasmid becomes stably integrated into the plant cell genome during infection. This property of the Ti plasmid has been exploited to show that DNA segments inserted in the T-DNA region can be cotransferred to plant DNA.

Applications

Recombinant DNA technology has permitted the isolation and detailed structural analysis of a large number of prokaryotic and eukaryotic genes. This contribution is especially significant in the eukaryotes because of their large genomes. The methods outlined above provide a means of fractionating and isolating individual genes, since each clone contains a single sequence or a few DNA sequences from a very large genome. Isolation of a particular sequence of interest has been facilitated by the ability to generate a large number of clones and to screen them with the appropriate “probe” (radioactively labeled RNA or DNA) molecules.

Genetic engineering techniques provide pure DNAs in amounts sufficient for mapping, sequencing, and direct structural analyses. Furthermore, gene structure-function relationships can be studied by reintroducing the cloned gene into a eukaryotic nucleus and assaying for transcriptional and translational activities. The DNA sequences can be altered by mutagenesis before their reintroduction in order to define precise functional regions.

Genetic engineering methodology has provided means for the large-scale production of polypeptides and proteins. It is now possible to produce a wide variety of foreign proteins in E. coli. These range from enzymes useful in molecular biology to a vast range of polypeptides with potential human therapeutic applications, such as insulin, interferon, growth hormone, immunoglobins, and enzymes involved in the dynamics of blood coagulation.