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Genetic Engineering

Genetic Engineering. Recombinant DNA & DNA Transformation. Recombinant DNA. New DNA (gene). Plasmid DNA. Restriction Enzymes “cut” Plasmid DNA Piece of DNA is Removed. New Piece (gene) of DNA is “stitched” to Plasmid DNA. Restriction Enzyme.

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Genetic Engineering

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  1. Genetic Engineering Recombinant DNA & DNA Transformation

  2. Recombinant DNA New DNA (gene) Plasmid DNA Restriction Enzymes “cut” Plasmid DNA Piece of DNA is Removed New Piece (gene) of DNA is “stitched” to Plasmid DNA

  3. Restriction Enzyme Bacterial Proteins that Cut Both Strands of the DNA Moloecules

  4. Plasmid DNA Small Ring of DNA Found in a Bacteria Cell E. Coli cell Plasmid DNA

  5. Applications of Recombinant DNA Bacteria Breaks Down Pollutants into Harmless Products Bacteria Extracts Minerals from Ores Insert the Human Gene into Bacteria to Produce Insulin for Diabetics Produce Artificial Sweeteners

  6. DNA Transformation 9th Grade Field Trip Museum of Science, Boston

  7. Objective: To transform the DNA of E. Coli bacteria by inserting a gene that will make the bacteria resistant to the antibiotic, ampicillin

  8. Ampicillin: A chemical that kills bacteria Plasmid: A circular piece of DNA found in bacteria pGREEN: A plasmid that contains genes that protects bacteria from ampicillin and makes the bacteria turn grenn LB broth: Food for bacteria LB plate: An Agar plate to grow bacteria on LB/Amp plate: An Agar plate that contains ampicillin Vocabulary

  9. Procedure at Museum of Science • 1. Use micro-pipets and innoculating sticks to mix calcium chloride solution with E. Coli bacteria. • 2. Label 4 Agar plates. • LB +pVIB • LB – pVIB • LB/Amp + pVIB • LB / Amp - pVIB • 3. Mix pVIB plasmid with appropiate bacteria / CaCl2 solution. 4. “Heat shock” bacteria in hot water bath and ice so that it takes in plasmid. 5. Spread + pVIB bacteria on “+”Agar plates 6. Spread – pVIB bacteria on “-” Agar plates 7. Incubate at room temperature for 72 hours and record bacterial growth.

  10. 1. Practicing sterilizing technique 2. New tip for micropipette 3. Preparing calcium chloride solution

  11. 4. Sterilizing inoculating stick 5. Scraping E. Coli bacteria 6. Adding E. Coli to CaCl2 solution

  12. 7. Inserting pVIB (plasmid DNA) into E. Coli 8. Inoculating Agar plates with genetically transformed E.Coli 9. Spreading bacteria evenly on Agar plates

  13. Agar Plates Before bacteria with gene antibiotics applied bacteria without gene antibiotics applied bacteria with gene normal growing conditions bacteria without gene normal growing conditions

  14. Agar Plates After 72 Hours - pVIB LB + pVIB LB bacteria without gene normal growing conditions bacteria with gene normal growing conditions + pVIB LB/ Amp - pVIB LB/ Amp bacteria without gene antibiotics applied bacteria with gene antibiotics applied

  15. Sample Results (2001)

  16. Questions to Ponder • What are the controls for this experiment? • Why is it important to practice sterilizing technique? • Did the students successfully insert the pVIB gene? • Why would you sometimes take antibiotics?

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