1. DNA TECHNOLOGY

2. DNA TECHNOLOGY • How can we use DNA? • Solve crimes, diagnose genetic disorders, solve paternity cases, compare species, study and cure disease, modify foods, etc. • Where do we get DNA samples? • Blood, skin, hair, semen, etc. • Human DNA • 99.9% the same; 0.1% different

3. Human Genome Project • Completed in 2003 • Map and sequence human genome • Find where all genes are located and sequence DNA • Uses: diagnose genetic disorders • Gene therapy • Insert normal genes into cells to correct genetic disorders

4. DNA Extraction • Open cells to isolate DNA from other cell parts

5. Polymerase Chain Reaction (PCR) • Used to amplify regions of DNA (make many copies of a small amount of DNA) • Heat DNA – to denature/unwind • Add primers (short segments of DNA) that match the gene you want to copy • Add polymerase and nucleotides • Repeat!

13. Restriction Enzymes • “cutting DNA” • Large DNA molecules are cut into smaller fragments • Restriction enzymes recognize and cut DNA at specific sequences

14. Restriction Enzymes • Come from bacteria • Ex: Eco RI • E=Escherichia(genus) • co=coli(species) • R=RY13(strain) • I=First identified - Order ID'd in bacterium • Cut DNA at specific point

15. Each Restriction Enzyme cuts DNA at a specific base sequence. For example, EcoRI always cuts DNA at GAATTC as indicated below. The sequence GAATTC appears three times in the DNA strand below. As a result, the strand is cut into four pieces.

16. Blunt Ends When RE cuts straight across DNA Sticky Ends Will join with other ends Restriction Enzymes AG CT TC GA GG CC CC GG G GATCC CCTAGG G A AGCTT TTCGA A G AATTC CTTAA G

17. Gel Electrophoresis • A way to separate DNA, RNA, or proteins based on size • Put sample in a gel and add electricity • DNA fragments move • DNA has negative charge (moves to + end) • Smaller fragments travel farthest • Bands are stained and compared to a standard to estimate size

22. Using Gel Electrophoresis • Solving a Crime • Compare the DNA bands • Must be an exact match • Paternity Test • Compare DNA bands • Child’s DNA must come from mother or father

25. DNA Fingerprinting • DNA sample is copied by PCR • DNA is cut into fragments by using restriction enzymes • Fragments of DNA are separated by gel electrophoresis • DNA is then compared for a “match”

26. Genetic Engineering • Genetic Engineering = Recombinant DNA Technology = Genetic Recombination • Cutting (cleaving) DNA from one organism into small fragments and inserting the fragments into a host organism of the same or different species • The host then uses the foreign DNA as its own

27. Genetic Engineering • Recombinant DNA = DNA fragments that have been connected (recombined) from different species • Transgenic Organsims = organisms that contain functional recombinant DNA (foreign DNA)

28. Genetic Engineering • How to Do It? • Isolate foreign DNA fragment • Attach DNA fragment to a “vehicle” • Transfer the vehicle into the host organism

29. Isolate Foreign DNA • Cut DNA from chromosome • Restriction Enzymes = bacterial proteins that have the ability to cut both strands of DNA at a certain sequence • Cut at palindromes • Blunt ends (cut straight across) • Sticky ends (leave single-stranded ends that can join with other ends)

30. Each Restriction Enzyme cuts DNA at a specific base sequence. For example, EcoRI always cuts DNA at GAATTC as indicated below. The sequence GAATTC appears three times in the DNA strand below. As a result, the strand is cut into four pieces.

31. Fragment Attached to Vehicle • Vector = used to carry DNA from one species to a host (can be biological or mechanical) • Plasmid = small ring of bacterial DNA • Micropipette • Gene Gun • Gene Splicing = rejoining DNA fragments • Cut DNA and plamsid with same restriction enzyme = ends match and join

32. Fragment Attached to Vehicle • If a plasmid and the foreign DNA are cut with the same restriction enzyme, the ends will attach and join • The recombinant DNA in the plasmid (vector) is put into a bacterial cell to clone the DNA

33. Uses of Recombinant Bacteria • Industry • To beak down pollutants • Clean up oil spills • Medicine • Make insulin, etc. • Transgenic organisms to study disease • Agriculture • Remove bacterial gene that causes frost in strawberries • Transgenic plants to resist herbicides or increase protein production