1. AP Biology 12 AP Biology 12 Concept 2: Analyzing and utilizing biotechnology tools Please refer to: Chapter 20 in Campbell Pg 136-141 in Holtzclaw Pg 307-313 in Holtzclaw Practice Questions: Campbell: #1-4 p. 405 (20.1) #1-3 p. 411 (20.2) #1-3 p. 416 (20.3) #1,4,5,6,7,8,9,10,11,12 p. 424-425 Holtzclaw: Questions from p. 143-150, 310, 312-313

2. Transgenic Organisms Transgenic Organisms This goat makes spider silk protein in its milk! How? Transgenic Organisms

3. Try This! One strand of a DNA molecule has the sequence 3 ′ -GGATGCCCTAGGCTTGTT-5 ′. Which of the following is the complementary strand? A.3 ′ -AACAAGCCTAGGGCATCC-5 ′ B.3 ′ -CCTACGGGATCCGAACAA-5 ′ C.5 ′ -AACAAGCCTAGGGCATCC-3 ′ D.5 ′ -GGATGCCCTAGGCTTGTT-3

4. Try This! One strand of a DNA molecule has the sequence 3 ′ -GGATGCCCTAGGCTTGTT-5 ′. Which of the following is the complementary strand? A.3 ′ -AACAAGCCTAGGGCATCC-5 ′ B.3 ′ -CCTACGGGATCCGAACAA-5 ′ C.5 ′ -AACAAGCCTAGGGCATCC-3 ′ D.5 ′ -GGATGCCCTAGGCTTGTT-3

5. Try This! You have isolated this eukaryotic gene and wish to express the protein it codes for in a culture of recombinant bacteria. Will you be able to produce a functioning protein with the gene as is? A.yes B.No, the exons will need to be cut out and the introns spliced back together. C.No, the introns will need to be cut out and the exons spliced back together. D.No, the exons will need to be cut out, the introns translated individually, and the peptides bound together after translation.

6. Try This! You have isolated this eukaryotic gene and wish to express the protein it codes for in a culture of recombinant bacteria. Will you be able to produce a functioning protein with the gene as is? A.yes B.No, the exons will need to be cut out and the introns spliced back together. C.No, the introns will need to be cut out and the exons spliced back together. D.No, the exons will need to be cut out, the introns translated individually, and the peptides bound together after translation.

7. We’ve come a long way! 1995 – first entire genome sequenced (bacteria) 2007 – only 12 years later ◦ Sequencing under way for 2000 organisms ◦ Complete human genome sequenced (3 billion base pairs!) ◦ Can look it up on the internetinternet

8. What are other examples of biotechnology feats? Biotechnology: manipulation of organism or their feats to make useful products ◦ Selective breeding (farms, dogs)  Choosing who breeds with who! ◦ Making wine and cheese and bread  Using bacteria/yeast! ◦ Genetic engineering  Making new proteins

9. So... What are the main techniques for manipulating DNA? Learning Intentions You must know: ◦ The terminology of biotechnology. ◦ The steps in gene cloning with special attention to the biotechnology tools that make cloning possible. ◦ The key ideas that make PCR possible ◦ How gel electrophoresis can be used to separate DNA fragments or protein molecules ◦ Examples of genetic engineering products

10. Learning Intentions for AP Investigation Labs – See Handout AP Investigation 7: Bacterial Transformation ◦ The principles of bacterial transformation, including how plasmids are engineered and taken up by cells ◦ Factors that affect transformation efficiency  Calculate transformation efficiency and express the results in scientific notation  Predict and justify how a change in the basic protocol for bacterial transformation would affect transformation efficiency ◦ How to verify and screen for transformed cells ◦ Bacterial transformation is a type of horizontal gene transfer, and increases genetic variation AP Investigation 9: Restriction Enzyme Analysis ◦ The function of restriction enzymes and their role in genetic engineering ◦ How gel electrophoresis separate DNA fragments ◦ How to use a standard curve to determine the size of DNA fragments  Apply mathematical routines to construct a graph of DNA fragments of known size  Use this standard curve to determine the size of unknown DNA fragments  Use the results of gel electrophoresis to map the restriction sites of a bacterial plasmid

11. Review: PCR and Electrophoresis http://learn.genetics.utah.edu/content/labs/pcr/ http://www.phschool.com/science/biology_place/labbench/lab6/intr o.html http://www.phschool.com/science/biology_place/labbench/lab6/intr o.html http://learn.genetics.utah.edu/content/labs/gel/ Complete: Investigation – How Can Gel Electrophoresis Be Used to Analyze DNA?

12. Try This! This segment of DNA is cut at restriction sites 1 and 2, which creates restriction fragments A, B, and C. Which of the following electrophoretic gels represents the separation of these fragments? a) b) c) d)

13. Let’s learn how to “clone” genes. Gene cloning: ◦ Process of producing multiple copies of specific DNA segments, making recombinant DNA in the process. ◦ Tools:  Restriction enzymes (Campbell – some one log into their student account please to show the class) – discovered in the 1960s by bacterial researchers Restriction enzymes

14. Cloning Genes Restriction enzymes ◦ How do they help bacteria in the wild?  Cut up foreign DNA (ex: phage DNA) ◦ How do they work?  Recognize restriction sites on DNA  Symmetrical/palindromal  4-8 nuclotide base pairs long  Make many cuts – produce restriction fragments  Cuts in a reproducible way  Bacterial DNA protected by methyl groups on restriction sites.

18. Try This! Which of the following DNA molecules is most likely to be cut by a restriction enzyme? A.5 ′ -AAGCCT-3 ′ B.5 ′ -GGAAGG-3 ′ C.5 ′ -GGATCC-3 ′ D.5 ′ -AATTAA-3 ′

19. Try This! Which of the following DNA molecules is most likely to be cut by a restriction enzyme? A.5 ′ -AAGCCT-3 ′ B.5 ′ -GGAAGG-3 ′ C.5 ′ -GGATCC-3 ′ (palindrome) D.5 ′ -AATTAA-3 ′

20. The process of Cloning genes Cloning a Gene in Bacteria Campbell Activity! Cloning a Gene in Bacteria

26. NOW… Transformation Lab! Handout – Lab Prepare by reading and making flow chart. INTENSE PROCEDURE!!! ◦ Make sure you understand it. Come into the lab knowing what you are doing. If you have questions, please ask!. ◦ Thurs, Jan 29 th ◦ Fri, Jan 30 th – lunch: Results

27. Also… You should now be able to answer all the questions of this section… Questions…