Genetic Technologies

1. Genetic Technologies Chapter 8

2. Genetic Engineering • Intent of altering human genome • Introducing new genetic material into genome • Insulin

3. Recombinant DNA • DNA that contains genes of two species • How? • Restriction enzymes – cut out desired gene • Occur naturally in prokaryotic cells • Recognize specific recognition sites – 4 to 8 base pairs • Recognition sites are palindromes • Cuts gene (digests) in one direction only • Creates restriction fragments

4. Process – Restriction Enzyme • Locates recognition site (Top Strand) • Cuts the DNA backbone • Locates recognition site (Bottom Strand) • Cuts the DNA backbone • DNA separates

5. Sticky Ends or Blunt Ends • Sticky ends – zigzag cuts in strand • Blunt ends – straight cut across strand

6. Putting Fragments together • DNA ligase – sticky ends • T4 DNIA ligase – blunt ends • forms phosphodiester bonds in DNA

7. Plasmids • Small circular pieces of DNA found in bacteria • Used as vectors for recombinant DNA (artificial) • Restriction enzymes used to isolate specific gene are used to cut plasmids

8. Process • Plasmids and DNA fragments are placed in same solution • Anneal • DNA ligase is used to form phosphdiester bond • Recombinant DNA introduced into host cell • DNA is cloned

10. Restriction Maps • Diagrams that show all recognition sites on a specific plasmid and distances in base pairs • Shows which restriction enzyme should be used • Allow scientists to determine which plasmids will work the best for cloning experiments

11. Transformation • Cells that receive foreign DNA • Bacterial cells sometimes will not take up a plasmid • Bacteria are placed in ice water bath containing CaCl₂ • Solution is heated and cooled repeatedly disrupting plasma membrane of bacteria allowing plasmid to enter • Solution is kept at 37⁰C to stabilize and grow

12. Identifying Bacteria Clones with Target Genes • Hybridization – identify cells that contain recombinant DNA • Identified using a hybridization probe – short single stranded complementary DNA molecule • Once identified bacteria can be grown in huge quantities (commercial use)

13. PCR • Polymerase Chain Reaction • Increase number of DNA copies from a single biological sample in a few hours • Only specific regions of a chromosome are replicated • Process • Denaturation • Annealing • Elongation

14. Taq polymerase is used to put strand together • Isolated from bacteria that live in hot springs

15. Gel Electrophoresis • Technique used to separate fragments of DNA (PCR)

16. Genetic Engineering • Biopharming • Pharmaceutical products produced on large scale • Organisms are genetically engineered to produce a specific protein • Ability to make new protein is passed on to offspring

17. Genetic Engineering • Transgenic Organism (genetically modified organism, GMO) • Organisms that contain one or more genes from another organism

18. Why use Genetic Engineering? • Cost • Larger organisms can produce larger molecules • Better versions of organisms • 80% of Canadas Canola crop is GM

19. Knockout Mice

20. Concerns?

21. Gene Therapy • Techniques used to replace, remove or alter a defective gene before symptoms are expressed

22. Gene Therapy • Germ-line gene therapy • Genes introduced in sperm or egg cells • Passed on to future generations • Somatic gene therapy • Genes introduced into body cells (not sperm or egg) • Will not be passed on

23. Genetic Screening • To detect any mutation in DNA that is associated with a genetic disorder (huntingtons disease) • Aminocentesis • Prenatal screening

24. Cord Blood (CBE) • Source of hematopoietic (blood forming) stem cells • Can be used to treat and cure disease and other conditions