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DNA TECHNOLOGY

DNA TECHNOLOGY. Genetic engineering and all that stuff. Glowing mice. Goals of Recombinant DNA Technology. Eliminate undesirable phenotypic traits in humans, animals, plants, and microbes Combine beneficial traits of two or more organisms to create valuable new organisms

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DNA TECHNOLOGY

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  1. DNA TECHNOLOGY Genetic engineering and all that stuff

  2. Glowing mice

  3. Goals of Recombinant DNA Technology • Eliminate undesirable phenotypic traits in humans, animals, plants, and microbes • Combine beneficial traits of two or more organisms to create valuable new organisms • Create organisms that synthesize products humans need

  4. Overview of recombinant DNA technology Figure 8.1.1

  5. Overview of recombinant DNA technology Figure 8.1.2

  6. Plasmid Vectors Example plasmid (pBR322)

  7. Tools and Techniques • Restriction Enzymes Cut DNA at precise locations based upon a specific target sequence of nucleotides • DNA Ligase Enzyme that “seals” or ligates DNA pieces together

  8. Restriction Endonucleases • Eco RI --GAATTC-- --G AATTC-- --CTTAAG-- --CTTAA G-- • SmaI --CCCGGG-- --CCC GGG-- --GGGCCC-- --GGG CCC-- Recognition sequenceResults of cutting

  9.   Using a restriction enzyme and DNA ligase to make recombinant DNA

  10. Cloning Steps • Isolate gene of interest (cut out of genome with a Restriction Enzyme) • Cut Vector with same R.E. to get compatible ends • Mix 2 DNAs together and Ligate(DNA Ligase) • Transform • Screen for successful event • Celebrate

  11. Cloning a human gene in a bacterial plasmid: a closer look

  12. Inserting DNA into Cells • Goal of DNA technology is insertion of DNA into cell (transformation) • Can use vectors and natural methods of transformation, transduction, and conjugation • Artificial methods • Electroporation • Protoplast fusion • Injection – gene gun and microinjection

  13. Inserting DNA into Cells

  14. Inserting DNA into Cells

  15. Tools and Techniques • Gel Electrophoresis

  16. Gel Electrophoresis Figure 8.7a

  17. Gel Electrophoresis of Macromolecules

  18. Forensic use of DNA patterns“DNA Fingerprints”

  19.  Using restriction fragment patterns to distinguish DNA from different alleles RFLPs restriction fragment length polymorphisms

  20.  DNA fingerprints from a murder case

  21. DNA Fingerprinting M = mother F = Father C = children both children show some bands with each parent Figure 8.13

  22. Tools and Techniques • Reverse transcriptase

  23.   Making complementary DNA (cDNA) for a eukaryotic gene

  24. Tools and Techniques • DNA synthesis • DNA sequencing

  25. DNA sequencers

  26. Tools and Techniques • PCR (Polymerase Chain Reaction) • Large number of identical molecules of DNA are produced in vitro • Allows any piece of DNA to be rapidly amplified ! • Only minute amounts of DNA are needed !

  27. PCR procedure • Heat to ~ 95 degrees to denature the DNA strands • Cool and elongate with heat tolerant DNA Polymerase (e.g. Taq polymerase from Thermus aquaticus) • Repeat cycle … Repeat cycle …Repeat cycle … Repeat cycle … Repeat cycle …

  28. Animation: Polymerase Chain Reaction PLAY

  29. Applications • Diagnosis of Disease • Gene Therapy • Vaccines and other Pharmaceuticals • Forensic use • Environmental use (bioremediation) • Agricultural use * Transgenic animals * Transgenic plants

  30. Transgenic organisms

  31.  One type of Gene Therapy

  32. PHARM animals Transgenic Sheep Carrying a human gene for a human protein which they secrete in their milk. The protein is then purified from their milk

  33. “Golden” rice contrasted with ordinary rice Transgenic Golden Rice (High levels of Beta-carotene –used to make Vitamin A – give the rice a golden color.

  34. Using the Ti plasmid as a vector for genetic engineering in plants TRANSGENIC PLANT

  35. Safety and Ethics GMO (genetically modified organism) food controversy

  36. Scare tactics?

  37. Consumer distrustJustified…silly…somewhere in between?

  38. BT Corn basics • BT protein is produced by the bacterium Bacillus thuringiensis. Toxic to some insects, nontoxic to all other life forms. • BT corn has the gene inserted so that the corn plant makes the BT protein. • Ingestion of BT protein by the larvae of the European Corn Borer kills the larvae • The corn plant now has it’s own defense

  39. Pluses and minuses • Reduced need for pesticides • Greater Yield, etc. etc. • Potential impact on other species None of any consequence noted…. …but still the GREAT UNKNOWN.

  40. The Biotech Floodgate 2008 Stats 90% of all soybeans 75% of all corn 65% of all foods on grocery store shelves (soy and corn syrup are in all sorts of stuff!)

  41. What’s Next • Bananas/potatoes that produce human vaccines • Fish that mature more quickly • Plants that produce plastics • Fruit/nut trees that yield years earlier • Crops that grow where they could not before • Human milk from cows • Etc. etc. etc. etc. etc. Two featherless chickens peck around in some grass at the Hebrew University in Rehovot. Israeli scientists at the Agriculture department of the university have genetically engineered bare-skinned chickens as part of a research project to develop succulent, low fat poultry that is environmentally friendly. (AP)

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