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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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