1. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings p.174

2. Biotechnology - use of living organisms to create products or help processes Ex. HGH, insulin Recombinant DNA - segment of DNA containing sequences from different organisms How is DNA manipulated?

3. A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A A T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T T C G C G C G C G C G C G C G C G C G C G C G G C C G C G C G C G C G C G C G G G C C C C G G C G C G G C G C G C G G GG G G G C C C C C Restriction enzymes cut DNA at specific sites and create sticky ends Complementary ends will fuse to produce a long strand of DNA

4. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings The DNA is then integrated into the recipient cell’s chromosome Figure 12.1D Donated DNA Recipient cell’s chromosome Crossovers Degraded DNA Recombinant chromosome

5. bacterium bacterial chromosome plasmid Plasmids are extra rings of DNA that replicate in bacteria. DNA can be inserted into plasmids.

6. Cloning Vectors

7. 1. Use restriction enzymes. 2. Insert gene into plasmid. 3. Transfer the plasmid back into bacterial cell. 4. Let bacterial cells replicate. bacterial clones replication transformation recombinant DNA Plasmid Bacterium Human cell DNA Human protein Bacterial chromosome

8. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Recombinant DNA products “seed” protein for artificial snow Insulin for diabetes treatment Enzymes that clean up toxic waste spills Growth Hormones (Human, Bovine) TPA: Tissue Plasminogen Activator for treatment of heart attacks

9. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings The polymerase chain reaction (PCR) can quickly clone a small sample of DNA in a test tube Selection of specific sequence Figure 12.12 Initial DNA segment 1248 Number of DNA molecules

10. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Restriction fragments of DNA are compared by size Gel electrophoresis sorts DNA molecules by size Figure 12.10 Mixture of DNA molecules of different sizes Power source Gel Glass plates Longer molecules Shorter molecules Completed gel

11. DNA forensics

12. Egg manipulation via microinjection. Credit: © Science VU/Visuals Unlimited Egg microinjection to produce transgenic animal

13. Grow bigger fish faster. Salmon with gene from another fish species

14. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Uses of transformed animals: Produce medicines Study human diseases Low-pollution pigs Low-fat pork Figure 12.16

15. An extremely large Agrobacterium tumefaciens tumor (crown gall disease) and secondary tumors on Kalanchoe stem. Credit: © Brad Mogen Genetic engineering of plants Methods to insert DNA: 1.Ballistics 2.Protoplasts 3.Agrobacterium as vector - Ti plasmid

16. a A bacterial cell contains a Ti plasmid (purple) that has a foreign gene (blue). b The bacterium infects a plant cell and transfers the Ti plasmid. The plasmid DNA becomes integrated into a plant chromosome. c The plant cell divides. Its descendant cells may develop into a plant that can express the foreign gene. A young plant expressing a fluorescent gene product Fig. 11-12, p.171

17. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Genetically modified crops Golden rice with Vitamin A Cotton resistant to boll weevil Soybeans resistant to herbicide (Roundup) Corn resistant to European corn borer Rapeseed with healthier vegetable oil Virus-resistant papaya Benefits and risks

19. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Gene therapy Goal - Treat diseases caused by mutated genes Method - Add a normal gene or block an abnormal gene in enough cells to restore normal function Target - somatic cells

20. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings treat disease by altering an afflicted individual’s genes –Ex vivo (shown here) –In vivo –Stem cells Gene therapy may someday help treat a variety of diseases Figure 12.19 Cloned gene (normal allele) 1 Insert normal gene into virus Viral nucleic acid Retrovirus 2 Infect bone marrow cell with virus 3 Viral DNA inserts into chromosome Bone marrow cell from patient Bone marrow 4 Inject cells into patient

21. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Which disorders are candidates for gene therapy treatment? Disorders due to mutations in one or more genes The responsible gene is known The affected tissues are known and accessible

22. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Knockout gene therapy Goal: turn off a gene that is causing a disorder Strategies: –Antisense –Triple helix oligos –Spliceosome –Ribozyme

23. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Critical factors in choosing a vector Gene size - limited room in vector genome Target tissue –What cells can the vector infect? Integration into the genome –Without integration, only short-term effect –Random integration may disrupt other genes Cell cycle stage (dividing vs. non-dividing)

24. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Gene Therapy Successes Ashanti de Silva successfully treated for ADA deficiency - 1990 Ryes Evans successfully treated for SCID - 2001 Photo courtesy of Van de Silva

25. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Gene Therapy Problems Jesse Gelsinger died of complications due to an immune system response while participating in a clinical trial Three children treated for SCID developed leukemia due to disruption of a gene that regulates cell division

26. Copyright © 2003 Pearson Education, Inc. publishing as Benjamin Cummings Ethical and Social Issues Patient safety while participating in clinical trials Which applications are therapies and which are enhancements? –“ Designer ” babies Access to gene therapies