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Agenda – Applying DNA knowledge to diabetes

Agenda – Applying DNA knowledge to diabetes

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Agenda – Applying DNA knowledge to diabetes

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  1. Agenda – Applying DNA knowledge to diabetes • Warm-up: • Attractive & professional thank-you notes • Diabetes, Insulin, and rDNA • Review • Recombinant DNA • Plasmids • Recombinant Paper Plasmid

  2. Recombinant DNA project • Notes on what you have learned – due tomorrow • Electronic presentations • http://prezi.com/ezf2bged4bia/genetic-engineering-and-biotechnology/Genetic Engineering and Biotechnology by Dong Min Kim on Prezi

  3. Diabetes and Biotechnology Biotechnology solutions Recombinant DNA

  4. Questions- • What is diabetes? • Why is there growing concern about diabetes? • What is the role of insulin? • What is the role of biotechnology in the treatment of diabetes?

  5. Biotechnology & Diabetes Treatment • Read Calorie-Coated Diabetes

  6. Beta Cell in Pancreas

  7. Pancreas, thymus

  8. Cell Biology Of Insulin Response

  9. Cell Biology Of Insulin Response

  10. Cell Biology Of Insulin Response

  11. Insulin Gene is on the 11th Chromosome (short arm) It contains 153 bases. A small, simple protein

  12. Recombinant DNA with insulin gene inserted Recombinant DNA: A desired gene to the plasmid. Bacteria is transformed to accept the plasmid.

  13. Chromosome & Plasmid

  14. Interesting facts about plasmids What do plasmids do in bacteria? Why are plasmids used in recombinant DNA? • In nature?

  15. Interesting facts about plasmids What do plasmids do in bacteria? Why are plasmids used in recombinant DNA? Bacteria can be transformed by adding circular DNA but not linear Exonuclease: Breaks down genes in a linear process Outside to the inside • Extra genes • Not essential for living • Benefit for survival under certain conditions In nature, some plasmids • Antibiotic genes • Antibiotic resistant genes

  16. Making a Recombinant DNA Product • What are the steps needed? • Paper plasmids - • Construction of the pAMP and pKAN Plasmids • Questions

  17. Genetic Engineering –Recombinant DNA How? • Identify a molecule produced by a living organism • Isolate the instructions (DNA sequence = gene) • Put the instructions into another cell or organism • Allow the cell to replicate • Harvest the desired product

  18. The Vector (E. coli bacteria).

  19. What is needed to produce a product with recombinant DNA?

  20. Recombinant DNA Insulin • Identify a insulin gene in humans • Use restriction enzymes to isolate the gene in a DNA fragment • DNA fragment is added to another DNA source = vector • such as plasmids of bacteria or yeast • Recombinant DNA is placed in a host cell • As the host cell divides (replicates), the rDNA also replicates • Harvest, purify, test & market

  21. What we need to learn about: • Bacteria (Vector) • Copying cells - Mitosis • Proteins • How are enough insulin is made? PCR

  22. Recombinant DNA & Cloning

  23. What is DNA cloning? • When DNA is extracted from an organism, all its genes are obtained • In gene (DNA) cloning a particular gene is copied (cloned)

  24. Why Clone DNA? A particular gene can be isolated and its nucleotide sequence determined Control sequences of DNA can be identified & analyzed Protein/enzyme/RNA function can be investigated Mutations can be identified e.g. gene defects related to specific diseases Organisms can be ‘engineered’ for specific purposes e.g. insulin production, insect resistance and more

  25. How is DNA cloned?, I Blood sample • DNA is extracted- here from blood • Restriction enzymes, e.g. EcoR I, Hind III, etc., cut the DNA into small pieces • Different DNA pieces cut with the same enzyme can join, or recombine. DNA Restriction enzymes

  26. DNA Cloning, II • Bacterial plasmids (small circular DNA additional to a bacteria’s regular DNA) are cut with the same restriction enzyme • A chunk of DNA can thus be inserted into the plasmid DNA to form a “recombinant” molecule

  27. DNA cloning, III • The recombinant plasmids are then mixed with bacteria which have been treated to make them “competent”, or capable of taking in the plasmids • This insertion is called transformation

  28. DNA Cloning, IV • The plasmids have naturally occurring (or inserted) genes for antibiotic resistance • Bacteria containing plasmids with these genes will grow on a medium containing the antibiotic- the others die, so only transformed bacteria survive

  29. DNA Cloning, V • The transformed bacterial cells form colonies on the medium • Each cell in a given colony has the same plasmid (& the same DNA) • Cells in different colonies have different plasmids (& different DNA fragments)

  30. Screening, I Screening can involve: • Phenotypic screening- the protein encoded by the gene changes the color of the colony • Using antibodies that recognize the protein produced by a particular gene

  31. Screening, II 3. Detecting the DNA sequence of a cloned gene with a probe (DNA hybridization)

  32. rDNA project • Choice of format: • PowerPoint Presentation with notes • Globster (electronic poster) • Museum boxes • Presentation: • Process to make rDNA • Process before rDNA • Benefits • Disadvantages or bioethical issues • Regulation • Equivalent to Test Grade

  33. Additional Resources http://www.accessexcellence.org/AE/AEC/CC/restriction.php Discovery and Applications of REs - http://videos.howstuffworks.com/hsw/22635-discoveries-with-bill-nye-restriction-enzymes-video.htm http://tools.neb.com/NEBcutter2/ http://www.accessexcellence.org/AE/AEC/CC/activity1.php