Lecture #9 Date _____

1. Lecture #9 Date _____ • Chapter 20~ DNA Technology & Genomics

2. Biotechnology • Biotechnology: manipulation of organisms or their components to perform practical tasks or provide useful products

3. Practical DNA Technology Uses • Diagnosis of disease • Human gene therapy • Pharmaceutical products (vaccines) http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter16/animations.html# • Forensics • Animal husbandry (transgenic organisms) • Genetic engineering in plants • Ethical concerns?

4. Selective breeding • Mating organisms to produce offspring with desired traits (has been around since ancient times) • Inbreeding= mating between closely related individuals • Hybrids– offspring of parents with different forms of traits. In plants, hybrids are often stronger or bigger.

5. Recombinant DNA • Recombinant DNA: DNA in which genes from 2 different sources are linked • Genetic engineering: direct manipulation of genes for practical purposes. (cutting out DNA from one organism and putting it into another.) (i.e. how we make recombinant DNA) • Transgenic organism= an organism with recombinant DNA • C:\Documents and Settings\BBAUGHMAN\Desktop\COURSES\AP BIO\articles • C:\Documents and Settings\BBAUGHMAN\Desktop\COURSES\LSSI Summer Institute Resources\LSSI Prog. Resources\GFP Mice

6. Genetic Engineering Tools • Restriction Enzymes • DNA ligase • Vectors • Plasmids! • Viruses • Reverse Transcriptase

7. Making recombinant DNA • Restriction enzymes : in nature, these enzymes protect bacteria from intruding DNA; they cut up the DNA (restriction) and are very specific • Restriction site: where the restriction enzyme cuts the DNA (a specific sequence) • Sticky end: short single-stranded extensions of restriction fragments (segments of DNA cut by restriction enzymes in a reproducible way)

8. Restriction Enzymes C:\Documents and Settings\BBAUGHMAN\Desktop\bio powerpoints\Chapter 13 BDOL IC http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter16/animations.html#

9. DNA ligase: enzyme that can join the sticky ends of DNA fragments (called gene splicing) • Vector: Something that carries foreign DNA into a cell. • Bacterial plasmids… small circles of DNA • viruses

10. Reverse Transcriptase • Catalyses the production of DNA from RNA • Why is this helpful in making recombinant DNA? • Hint: Think about introns and exons… • Ex. Bacteria that make human insulin…

11. RNA viruses • Where does Reverse Transcriptase come from? • Answer: • Retroviruses: transcribe DNA from an RNA template (RNA--->DNA) • DNA gets incorporated as provirus in host • Ex. HIV • http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter26/animations.html • http://www.pbs.org/wgbh/nova/aids/action.html

12. Bacterial plasmids in gene cloning

13. Gene Cloning • Clone = genetically identical copy • Gene cloning: Making more copies of a gene • Bacterial plasmid with recombinant DNA reproduces as the bacteria reproduce. • C:\Documents and Settings\BBAUGHMAN\Desktop\bio powerpoints\Chapter 13 BDOL IC

14. Steps for eukaryotic gene cloning • Isolation of cloning vector (bacterial plasmid) & DNA (gene of interest) • Insertion of gene of interest into the cloning vector (how?) • Introduction of cloning vector into cells (transformation by bacterial cells) • Cloning of cells (and foreign genes) • Identification of cell clones carrying the gene of interest • (nucleic acid hybridization– use radioactively labeled nucleic acid probe) or • look for protein product http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter16/animations.html#

15. Examples of GMO’s • Beta-carotene rice (a.k.a. Golden rice, vitamin A rice)-- daffodil and soil bacterium genes • Bt corn and other crops— Bacillus thuringiensis gene. • Terminator technologies– cause sterility of seeds.

16. Gene Therapy Germ line Therapy: reproductive cells– affects offspring Somatic Therapy: body cells of an individual Ex. White blood cells of SCID (severe combined immunodeficiency) Blood cells are removed Use of viral vector Blood cells replaced, having needed gene. Risks?

17. Animal Cloning • Clone= a group of genetically identical organisms or a group of cells derived from a single parent cell. • ex. Dolly the sheep • Somatic cell nuclear transfer • Remove Nucleus from an egg • Fuse this unfertilized egg with a body cell • Ethical issues • Reproductive cloning • Therapeutic cloning– creation of an embryo to provide stem cells • http://videos.howstuffworks.com/hsw/11818-genetics-cloning-video.htm

19. DNA Analysis & Genomics • PCR (polymerase chain reaction) • Gel electrophoresis • Restriction fragment analysis (RFLPs) • DNA sequencing • Human genomeproject

20. Polymerase chain reaction (PCR) • Amplification of any piece of DNA without cells (in vitro) • Materials: heat, primers (single-stranded DNA), DNA polymerase, nucleotides, • Applications: fossils, forensics, prenatal diagnosis, etc. • http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter16/animations.html#

21. DNA Analysis • Gel electrophoresis: separates nucleic acids or proteins on the basis of size or electrical charge creating bands of DNA consisting of pieces of the same length. • Used in DNA Profiling • Paternity tests • Forensics • Ethical issues? • Good DNA ANALYSIS COMPUTER LAB ACTIVITY= http://www.biology.arizona.edu/human_bio/activities/blackett/introduction.html • See Activities for in class simulation to be used for forensics and paternity tests

22. How does gel electrophoresis work? • DNA is cut with restriction enzymes and added to a well. • The negative DNA is attracted to the positive electrode. • smaller pieces of DNA move faster through the pores in the gel • A unique banding pattern is observed. • The human genome is estimated to be about 3 billion base pairs long and to contain 20,000-25,000 distinct genes

23. Gel Electrophoresis animationhttp://www.sumanasinc.com/webcontent/anisamples/majorsbiology/gelelectrophoresis.html http://www.dnalc.org/ddnalc/resources/animations.html http://learn.genetics.utah.edu/units/biotech/gel/ good gel electrophoresis virtual lab

24. DNA Analysis (continued) • Restriction fragment length polymorphisms (RFLPs)= differences between different individuals in lengths of restriction fragments • Found especially in satellite DNA (highly repetitive non-coding sequences of DNA) • RFLP animation http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter16/animations.html# • Southern blotting: process that reveals sequences and the RFLPs in a DNA sequence http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter16/animations.html#

25. O.J. Simpson capital murder case,1/95-9/95 • Odds of blood in Ford Bronco not being R. Goldman’s: • 6.5 billion to 1 • Odds of blood on socks in bedroom not being N. Brown-Simpson’s: • 8.5 billion to 1 • Odds of blood on glove not being from R. Goldman, N. Brown-Simpson, and O.J. Simpson: • 21.5 billion to 1 • Number of people on planet earth: • 6.1 billion • Odds of being struck by lightning in the U.S.: • 2.8 million to 1 • Odds of winning the Illinois Big Game lottery: • 76 million to 1 • Odds of getting killed driving to the gas station to buy a lottery ticket • 4.5 million to 1 • Odds of seeing 3 albino deer at the same time: • 85 million to 1 • Odds of having quintuplets: • 85 million to 1 • Odds of being struck by a meteorite: • 10 trillion to 1

26. DNA Sequencing • Determination of nucleotide sequences • Genomics: the study of genomes based on DNA sequences • Genome: all of an organism’s genetic material. • Human Genome Project Animations: http://www.genome.gov/25019885

27. Human Genome Project • Outcomes: • Identification and sequencing of genes involved in traits and disease. • Genetic testing improved. • Greater understanding of genetics. • Ethical issues with genetic testing etc.\ http://www.pbs.org/wgbh/nova/sciencenow/archive/title-m-z.html Public Genomes or Personal genetic Testing