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| 19.1 Of Pigs and Patents 1. A transgenic organism contains foreign DNA. 2. Biotechnology is the industry dedicated to using altered cells or molecules for various applications. 3. Recombinant DNA technology (i.e. gene cloning) was the first of the modern biotechnologies. 4. Gene targeting uses precision techniques to "knockout" a gene or substitute a gene for one on a chromosome. 5. Bioethics questions, gene patenting, and potential uses and risks of biotechnology pose treat questions for society. 19.2 Recombinant DNA TechnologyConstructing Recombinant DNA Molecules -- An Overview 1. Recombinant DNA molecules are made by covalently linking foreign DNA with the DNA of a transport molecule known as a cloning vector (i.e. plasmid DNA). 2. Recombinant DNA is amplified by introducing it into a bacterium or other single cell host. 3. Guidelines for using recombinant DNA technology were first drawn up at a 1975 meeting at Asilomar, CA. Isolating the Gene of Interest 1. Bacteria and other single cell organisms are also used as hosts of DNA libraries. 2. DNA libraries can be constructed from the DNA fragments of chromosomes (genomic libraries) or can contain only processed coding sequences when mRNA is used as the source of the genetic information (cDNA libraries). 3. DNA probes are used to identify and isolate specific DNA fragments of interest from DNA libraries. Selecting Recombinant DNA Molecules 1. Phenotype selection can be used to identify host cells with recombinant DNA. 2. A popular technique identifies host cells with recombinant DNA using antibiotics and a color test. Delivering DNA in Plants and Animals 1. Methods of gene transfer include Ti plasmid in dicots and other methods in monocots. 2. Genes in the nucleus or chloroplast can be manipulated. 3. Many crop species have been engineered to be herbicide or pesticide resistant. 4. The nutritional value of numerous crops has been enhanced. 5. Root systems of engineered plants, through rhizosecretion, can be used for pharmaceutical production. 6. Altering the levels of gene products (promotor enhancement; antisense knockouts) has been widely applied. 19.3 Applications of Recombinant DNA Technology 1. The products of recombinant DNA technology are used in health care, food technology, agriculture, and forensics. Drugs 1. The first application of recombinant DNA technology was to mass produce human insulin, a protein-based drug. Textiles 1. Recombinant DNA technology has created a new source of indigo, the dye used to make blue jeans blue. Paper and Wood Products 1. Several popular species are emerging as valuable model systems for experimenting with transgenic trees. Food 1. Ti plasmids have helped to develop novel, or easier-to-grow fruits and vegetables in countries that do not restrict the marketing of genetically modified foods. Bioremediation 1. Bioremediation is the intentional release of organisms that detoxify certain pollutants. 2. The enzyme-catalyzed detoxification reactions may be a natural part of an organism's biochemistry or may be added transgenically. 3. Organisms used in bioremediation help to detect or detoxify heavy metals, organic wastes, or hidden explosives (land mines) in the environment. 19.4 Gene Targeting 1. In contrast to transgenic organisms, where DNA is randomly inserted into chromosomes, gene targeting in mice involves homologous recombination and the controlled insertion of a gene at a specific chromosomal locus an embryonic stem (ES) cell. 2. The ES cell is then incorporated into a developing embryo from another individual, which is implanted into a surrogate. Gene Targeted Mice as Models 1. Animals with phenotypes indicating that they harbor cells with the targeted gene are bred to each other to yield homozygotes for the targeted gene. 2. Swapping an inactivated allele for a gene of interest produces a knockout mouse, and replacing a gene with another that has an altered function creates a knockin mouse. 3. Knockout mice can often serve as useful animal models for human diseases. 4. Sometimes animals with supposedly vital genes knocked out are perfectly healthy and the importance of the gene product must then be reconsidered at the level of the entire organism. 19.5 Antisense Technology 1. Antisense technology inserts a DNA sequence in revese orientation so that the "antisense" mRNA transcribed from its complementary base pairs with the "sense" mRNA, squelching a gene's expression. 2. Limitations have slowed its progress, but products are still being tested. |