Recombinant DNA Technology Help
Recombinant DNA Technology
Two primary advances helped usher in the new era of molecular genetics in the late 1970s to the early 1980s: (1) the use of recombinant DNA technology (genetic engineering) used to isolate and manipulate genes in vitro in order to endow cells with new synthetic capabilities and (2) the ability to synthesize and determine the linear order of nucleotides of DNA molecules (DNA sequencing). Recombinant DNA technology has made it possible to clone (isolate and make copies of individual genes) and transfer genes between bacterial species and strains or from eukaryotes into bacteria (or vice versa), causing the engineered cells to produce, sometimes in relatively large quantities, proteins of great economic importance such as enzymes (e. g. , amylases, proteases), hormones (e. g. , insulin, growth hormone), and interferons (lymphocyte proteins that prevent replication of many viruses). These proteins are made in such small quantities in human cells that the cost of their extraction and purification from blood or cadaver tissues has been very expensive, thus restricting their medical use in prevention and treatment of disease. Many products are now produced successfully using genetic engineering techniques; such products include blood-clotting factors (e. g. , tissue plasminogen activator, or TPA, used to activate the breakdown of blood clots and to prevent recurrence of blood clots in heart attack patients), and complement components (part of the immune system). In 1980, the United States Supreme Court decreed that new life forms created by genetic engineering could be patented, although this is not true in many other countries. This decision has contributed to the investment of large sums of money by private corporations into the development of many useful recombinant strategies. Public outcry against recombinant, or GM (genetically modified), crops in Europe had led the European Union (E. U. ) to ban or require extensive labeling requirements of foods that contain ingredients from GM crops. This has caused significant tension between the U. S. food industry and E. U. countries.
There are many hopes for the future benefit of genetic engineering technology in helping solve some of society's problems. In the agricultural arena, soybeans, corn, and cotton have been genetically engineered or modified to contain added herbicide tolerance and pest-resistance genes. These crops are also known asGMOs (genetically modified organisms). For example, one type of GM corn (Bt corn) contains a gene from the bacterium Bacillus thuringiensis that produces a protein that kills insect larvae. Larvae of the European corn borer can be costly pests for corn farmers. Having the pesticide gene in the corn eliminates the need to spray the crop with a pesticide. Other crops have been or are being engineered to contain genes that encode vaccines, added nutrients, and disease resistance, as well as resistance against cold, drought, and salinity. These few examples suffice to demonstrate the possibilities of this new technology and explain why there is such great excitement in the scientific, medical, agricultural, and pharmaceutical communities concerning its further development. While this technology is exciting and has proven useful, some have voiced concerns over potential environmental hazards, unknown risks to human health, and economic impact. These issues are still being investigated and must be resolved in order to make this technology useful and safe.
Most genetic engineering projects begin with the cloning, or isolation and amplification, of a gene. There are two major purposes for cloning a gene: (1) to obtain the sequence of its DNA (e. g. , in order to discover how an abnormal gene differs structurally from its normal allele) and (2) to produce large amounts of a gene product (i. e. , protein). Gene cloning generally involves three major steps: (1) A DNA fragment (or gene) of interest is isolated. (2) The fragment is spliced into a cloning vehicle called a cloning vector, such as a phage or a plasmid. (3) The vector is introduced into a host cell where it is replicated many times using the host cell's DNA replication machinery. A good cloning vector should have three characteristics: (1) the ability to replicate autonomously from the main chromosome to make many copies within the cell, (2) one or more unique recognition sites for restriction enzymes so it can be opened at a single position to receive a foreign DNA insert (i. e. , the gene of interest being cloned), and (3) a gene that encodes resistance to an antibiotic agent that will allow for the distinguishing of cells that contain the cloning vector as opposed to cells that do not.
Once the gene is cloned it can be analyzed (restriction mapped and its DNA sequence determined) and then transferred into a host cell or organism that will allow it to be expressed as a protein product. The host system might be a genetically engineered (transgenic) plant or a mammalian cell culture, yeast culture, or bacterial cell system that is used as a biological factory to produce large quantities of the desired product.
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