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Chapter 9

Chapter 9. Biotechnology and Genetic Engineering . Biotechnology and Recombinant DNA. Biotechnology Genetic engineering Transgenic organisms (GMOs) “biological factories” Uses recombinant DNA techniques inserts foreign DNA to modify existing DNA sequence. Requires vector

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Chapter 9

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  1. Chapter 9 Biotechnology and Genetic Engineering

  2. Biotechnology and Recombinant DNA • Biotechnology • Genetic engineering • Transgenic organisms (GMOs) • “biological factories” • Uses recombinant DNA techniques • inserts foreign DNA to modify existing DNA sequence

  3. Requires vector • Produces clones carrying gene of interest Figure 9.1.1

  4. Genetic Engineering Using Agrobacterium

  5. Recombinant DNA Procedures • Protoplast fusion • Electroporation • Gene gun • Microinjection

  6. Protoplast Fusion

  7. Electroporation Microinjection Gene gun

  8. Restriction enzymes • Naturally occurring bacterial enzymes • Cut DNA in predictable and controllable manner • Generates restriction fragments • Covalently joined with ligase • Thousands of restriction enzymes • Specific target sites • Used to excise genes of interest

  9. Gene Amplification • The Polymerase Chain Reaction (PCR) • Amplifies small amounts of DNA • Creates millions of copies of DNA fragment in matter of hours • Many cycles (30-35) of the following steps: • Denaturing • Annealing • Synthesis

  10. Components of PCR Reaction • Template (DNA Fragment) • Primers • Deoxyribonucleotides • DNA polymerase • Heat tolerant Taq Polymerase isolated from Thermus aquaticus • Cycles are controlled by temperature alterations • thermalcycler

  11. Denature Anneal Synthesis

  12. Gel electrophoresis • Separates DNA fragments according to size • DNA put into wells in gel • Gel subjected to current • DNA moves through the gel • Fragments separated according to size

  13. Sources of DNA 1.Gene libraries • Extract total DNA from cells and digest with restriction enzymes • Create recombinant vectors to generate clones

  14. Eukaryotic genes require cDNA made from mRNA with reverse transcriptase

  15. 2. Synthetic DNA made with a DNA synthesis machine • Exact sequence of gene must be known

  16. Applications of Recombinant DNA • Agricultural • Pest and herbicide resistance • Improved nutritional value • Pharmaceutical proteins added to animal and plant products

  17. Medical • Pharmaceutical proteins • Vaccine production • Gene therapy

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