1. What Is Biotechnology? Any technique that uses living organisms or substances from those organisms to make or modify a product improve plants or animals to develop microorganisms for specific uses

2. WHAT COULD SOME POSSIBLE REASONS BE???? Adding a clotting agent to the blood that would help dissolve clots. TPA--- DONE!!!!! What if: Cancer free genetics HIV free genetics Creating something that was resistant to these diseases??? IS IT POSSIBLE?

3. Genetic Engineering Genetic engineering is taking one or more genes from one organism and ….. Transferring them to another organism Putting them back into the original organism in different combinations GMO’s: Transgenic Organisms: Bt Corn ( pest resistant) Recombinant DNA (rDNA): rInsulin

4. What are the Benefits of Biotechnology? Medicine Human TPA, Dissolve blood clots Cancer Treatments Embryo Screening Veterinary Hybrid Dogs Surrogate Cows Environment Clean up oil spills (bioremediation) Agriculture Bt Corn ( pest resistant) Food products Cheese, Yogurt, Fermented foods, Bread, Alcohol) Industry/manufacturing Better cleaners

5. rInsulin The first commercial product made by genetic engineering (1976). Insulin is the hormone used to control their blood sugar levels in diabetics First sold in 1982, (S. San Francisco) PIGS E. Coli

6. Biotech Tools Restriction Enzyme Restriction Enzyme: an enzyme that cuts DNA Ligase Ligase: an enzyme that pastes DNA back together  re-establishes the ester bond Gene of Interest Gene of Interest: Example Insulin Gene Recipient Genome Recipient Genome: E. coli plasmid  Plasmid: Circular DNA that bacteria have in addition to the normal chromosome

7. Restriction Enzyme Natural function of restriction enzymes To protect bacteria against viruses. Digest foreign (e.g. viral) DNA. Restriction enzymes are isolated from bacteria Restriction Enzyme Restriction Enzyme: an enzyme that cuts DNA

8. A closer look…. EroR1 5’…. ACTGTACGAATTCGCTA….3’ 3’….TGACATGCTTAAGCGAT…. 5’ EcoR1

9. A closer look…. EcoR1 AATTCGCTA….3’ GCGAT….5’ 5’….ACTGTACG 3’….TGACATGCTTAA “sticky ends” -can bind with other DNA molecules with the same overhangs

10. 5’….ACTGTACAGATCCGCTA….3’ 3’….TGACATGTCTAGGCGAT….5’ DNA Ligase DNA ligase

11. Create Your Own Recombinant DNA ! 1. Using the shorter sequence, create your plasmid, remember they are circular 2. Tape= DNA Ligase ; Scissors= restriction enzymes 3. Highlight the gene of interest TAATGGATCCTT 4. Figure our what restriction enzyme to use. GAATTC EcoR1 Sequence CTTAAG GGATCC BamH1 Sequence CCTAGG 4. The plasmid and DNA must be cut with same Restriction Enzyme 5. Create your Recombinant DNA.

13. HOW IS IT DONE? Steps for genetically engineering Insulin Steps for genetically engineering Insulin 1. Take a human cell and remove the nucleus 2. Extract the DNA (genome) from the nucleus 3. Locate the Insulin gene 4. Cut out the Insulin gene with a restriction enzyme 5. Take a bacterial plasmid and cut open with the same restriction enzyme 6. Place the Insulin gene into the bacterial plasmid; you now have recombinant DNA. 7. Place the recombinant DNA back into an bacterial cell (transformation) 8. The bacteria will now produce insulin.

14. A closer look…. HindIII 5’…. ACTGTACAAGCTTGCTA….3’ 3’….TGACATGTTCGAACGAT…. 5’ HindIII

15. A closer look…. HindIII AGCTTGCTA….3’ ACGAT….5’ 5’….ACTGTACA 3’….TGACATGTTCGA “sticky ends” -can bind with other DNA molecules with the same overhangs

16. Restriction Enzymes & DNA Evidence Original Cases Timeline 1984 Alec Jeffreys develops DNA fingerprinting, for forensics and paternity cases. 1987 Tommie Lee Andrews is convicted of rape in Orlando based on DNA fingerprinting--the first such case in the nation. 1989 DNA evidence for the first time overturns a conviction, clearing Gary Dotson of an Illinois rape.

17. Famous DNA Cases The Romanov's and Anastasia OJ Simpson (DNA proved guilt but people didn’t understand DNA evidence) Thomas Jefferson and Sallie Heming's kids Use to identify 9-11 remains Innocence Project : to date 305 wrongful convictions overturned by DNA evidence, that wasn’t available at the time

18. Gel electrophoresis Cuts DNA at repetitive DNA that everyone has but are unique called Restriction Fragment Length Polymorphisms (RFLP) We can use the fragments made by the restriction enzyme digestion to make a DNA fingerprint Looking for ~13 variations DNA is negatively charged (due to phosphate backbone) When we pump electrical charge into the buffer, it will make the DNA move towards the positive charge

19. Imagine you are a DNA molecule Sorts DNA by length Smaller DNA fragments travel further in the gel If you were inside an agarose gel, your environment would resemble a very dense web. The smaller the DNA fragment, the easier it is to get through the web. This creates a unique DNA banding pattern called a DNA Fingerprint http://www.sumanasinc.com/we bcontent/animations/content/gel electrophoresis.html

20. Crime Scene DNA Who Committed The Crime?

21. Famous DNA Case

22. Let’s Try it Crime Scene DNA GAATTCTAATGGATCCTT G AATTC EcoR1 Sequence G GATCC BamH1 Sequence Creates 3 pieces 1 bp, 10 bp, 7 bp 10 bp 7 bp 1 bp + _

23. Lab: Crime Scene Analysis 1. Restriction digestion with BamH1 (GGATCC) Use blue pen 2. Restriction digestion with EcoR1 (GAATTC) Use red pen 3. Plot fragments on mock gel (based on fragment sizes) 4. Exclude suspects 5. Answer questions on mock gel WS 6. Answer wrap questions 7. Find news article on DNA fingerprinting and summarize article

26. Somatic Cell Nuclear Transfer (SCNT) aka: Cloning 1952 – Xenopus Tadpoles 1996 – The first mammal cloned from adult cells was Dolly, the sheep. 2001 – The first human embryo cloned, but on divided to six cells, providing evidence that human cloning is not currently possible http://www.youtube.com/watch?v=bbZiOiPVG6c

27. Dolly: 277 tries= 1 Sheep On 14 February 2003, Dolly was euthanized because she had a progressive lung disease and severe arthritis. Normal Life expectancy: 11-12 yrs.

28. Let’s Click and Clone http://learn.genetics.utah.edu/content/tech/cl oning/clickandclone/ http://learn.genetics.utah.edu/content/tech/cl oning/clickandclone/