Genetic Modification and Biotechnology

Genetic Modification and Biotechnology Experimental Techniques Describe the purpose and process of the polymerase chain reaction (PCR) PCR is used to rapidly amplify minute quantities of DNA ……………………………………………………………………………………………………… It involves a thermal cycler and three repeating steps: ……………………………………………………………………………………………………… • Denaturation - DNA is heated to separate strands ……………………………………………………………………………………………………… • Annealing – Primers are introduced to designate copying points ……………………………………………………………………………………………………… • Elongation - Taq polymerase* synthesises new strand ……………………………………………………………………………………………………… These three steps double the amount of DNA, so a typical reaction of ……………………………………………………………………………………………………… 30 cycles will produce over 1 billion copies of desired DNA sequence ……………………………………………………………………………………………………… * Taq polymerase is heat resistant and so doesn’t denature during PCR ……………………………………………………………………………………………………… Describe the purpose and process of gel electrophoresis (with regards to both DNA and proteins) Gel electrophoresis separates fragments of DNA or protein based on size ………………………………………………………………………………………………………………………………………………….......... DNA is run in an agarose gel (moves towards the positive terminus as phosphate is negatively charged) ………………………………………………………………………………………………………………………………………………….......... Proteins are run on a polyacrylamide gel after first being treated with SDS (homogenises protein charge) ………………………………………………………………………………………………………………………………………………….......... ………………………………………………………………………………………………………………………………………………….......... On the following diagram, identify the terminals and order the fragments from largest to smallest Larger fragments move ………………………………………… 5 3 – 1 2 4 more slowly through the ………………………………………… + gel matrix and hence do ………………………………………… not travel as far ………………………………………… ………………………………………… Identify two types of vectors for gene transfer Plasmids 1. …………………………………………………………………………………………………………………………………………………... Viruses 2. ………………………………………………………………………………………………………………………………………………….. Distinguish between ‘blunt’ and ‘sticky end’ restriction endonucleases Sticky ends are overhanging sequences which allow for specificity in ligation (only complementary ………………………………………………………………………………………………………………………………………………….......... sequences can ligate together) – blunt ends have no overhang ………………………………………………………………………………………………………………………………………………….......... With the aid of the diagram, outline the process of gene transfer Gene of interest and plasmid vector are isolated from cells and amplified with PCR ………………………………………………………………………………………………………………………………………………….......... Gene and plasmid are cut with restriction enzymes ………………………………………………………………………………………………………………………………………………….......... Gene and plasmid are fused together by DNA ligase to form a recombinant construct ………………………………………………………………………………………………………………………………………………….......... The construct is then inserted into a host cell (via any of a number of different techniques) ………………………………………………………………………………………………………………………………………………….......... Transgenic cells are then selected by growing on antibiotic media (the plasmid has a resistance gene) ………………………………………………………………………………………………………………………………………………….......... The transgenic cells are then grown in culture and the newly expressed protein can then be extracted ………………………………………………………………………………………………………………………………………………….......... An example of this process is the mass production of human insulin by transgenic bacteria ………………………………………………………………………………………………………………………………………………….......... ………………………………………………………………………………………………………………………………………………….......... Experimental Applications Explain the role short tandem repeats (STRs) play in DNA profiling STRs are short repeating sequences in non-coding DNA ………………………………………………………………………………………………………………………………………………….......... Different individuals have different numbers of repeats, so by comparing STR loci, unique profiles emerge ………………………………………………………………………………………………………………………………………………….......... ………………………………………………………………………………………………………………………………………………….......... Outline two potential applications for DNA profiling Forensic investigations ………………………………………………………………………………………………………………………………………………….......... Paternity testing ………………………………………………………………………………………………………………………………………………….......... Analyse the following DNA profile to determine paternity Stevensons Baby 1: …………………………………………………………………………………………………………………………………………... Jones Baby 2: …………………………………………………………………………………………………………………………………………... Smith Baby 3: …………………………………………………………………………………………………………………………………………... Outline the benefits and risks associated with the use of genetically modified crops Benefits Risks ………………………………………………………………………… Risk of cross-contamination (i.e. super weeds) ………………………………………………………………………… Improve yields Potentially longer shelf life ………………………………………………………………………… Reduce need for chemical pesticides ………………………………………………………………………… Allows growth in wider areas (e.g. salty soils) Threatens native biodiversity ………………………………………………………………………… Difficult to police internationally ………………………………………………………………………… Patenting could lead to monopolies ………………………………………………………………………… ………………………………………………………………………… ………………………………………………………………………… ………………………………………………………………………… ………………………………………………………………………… ………………………………………………………………………… Cloning Define clone Groups of genetically identical cells or organisms ………………………………………………………………………………………………………………………………………………….......... Identify four methods of natural cloning in animals Budding - e.g. yeast 1. …………………………………………………………………………………………………………………………………………………... Binary fission - e.g. Hydra 2. ………………………………………………………………………………………………………………………………………………….. Fragmentation - e.g. starfish 3. …………………………………………………………………………………………………………………………………………………... Parthenogenesis - e.g. some fish species 4. ………………………………………………………………………………………………………………………………………………….. Outline how stem cuttings can be used to clone plants Plants retain pluripotent cells (called meristem) and can reproduce asexually via vegetative propagation ………………………………………………………………………………………………………………………………………………….......... Hence new plant clones can be grown from stem cuttings ………………………………………………………………………………………………………………………………………………….......... With the aid of the following diagrams, explain how organisms can be cloned artificially Embryo Division Somatic Cell Nuclear Transfer (SCNT) Embryonic cells retain pluripotency Nucleus removed from diploid body cell of an adult By dividing the embryo into two cell masses, Egg is enucleated via UV radiation exposure each can grow into a new organism Diploid nucleus is inserted into enucleated egg Because these cell masses have identical Egg stimulated to divide by electric shock genetic information, they will form clones As all DNA came from diploid nucleus, the resulting ……………………………………………………………………… ……………………………………………………………………………. ……………………………………………………………………… ……………………………………………………………………………. ……………………………………………………………………… ……………………………………………………………………………. ……………………………………………………………………… ……………………………………………………………………………. ……………………………………………………………………… ……………………………………………………………………………. embryo will be a clone of the adult ……………………………………………………………………… …………………………………………………………………………….