(See related pages)
|
|
|
| 18.1 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. 18.2 Recombinant DNA TechnologyConstructing Recombinant DNA Molecules 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. 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. 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. Applications of Recombinant DNA Technology 1. The products of recombinant DNA technology are used in health care, food technology, agriculture, and forensics. 18.3 Transgenic OrganismsDelivering DNA 1. Introducing foreign DNA into a single germline cell creates a transgenic organism. 2. Entry of foreign DNA into cells requires the use of chemical agents (calcium phosphate, liposomes), physical methods (electroporation, microinjection, particle bombardment), and biological vehicles (viruses or Agrobacterium). 3. The foreign DNA is then transmitted to each cell as the organism develops. Transgenic Pharming from Milk and Semen 1. Animals are transgenically altered to "pharm" useful human proteins in their milk or germline cells (eggs or sperm). 2. Transgenic animals are also valuable research tools as models of human disease. 18.4 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. When Knockouts Are Normal 1. Knockout mice can often serve as useful animal models for human diseases. 2. 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. |