1. 1 Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Human Biology Sylvia S. Mader Michael Windelspecht Chapter 21 DNA Biology and Technology Lecture Outline Part 3

2. 2 Regulation of gene expression 5 levels 1. ____________________ (nucleus) e.g., chromatin density and DNA accessibility 2. Transcriptional control (nucleus) e.g., transcription factors 3. Posttranscriptional control (nucleus) e.g., mRNA processing 4. Translational control (cytoplasm) e.g., differential ability of mRNA to bind ribosomes 5. Posttranslational control (cytoplasm) e.g., changes to the protein to make it functional 21.2 Gene Expression

3. 3 Regulation of gene expression Figure 21.13 Control of gene expression in eukaryotic cells. 21.2 Gene Expression Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Cytoplasm Nucleus nucleosome signal chromatin packing DNA DNA unpacking mRNA processing mRNA translation Prosttranslational control primary mRNA maturem RNA nuclear envelope nuclear pore polypeptide functional protein degraded protein DNA transcription exonintron

4. 4 DNA technology 1.Gene cloning through recombinant DNA 2.Polymerase chain reaction (PCR) 3.DNA fingerprinting 4.Biotechnology products from bacteria, plants, and animals 21.3 DNA Technology

5. 5 DNA technology terms ______________________ – altering DNA in bacteria, viruses, plants, and animal cells through recombinant DNA technology Recombinant DNA – contains DNA from 2 or more different sources Transgenic organisms – organisms that have a ________ gene inserted into them Biotechnology – using natural biological systems to create a product or to achieve an end desired by humans 21.3 DNA Technology

6. 6 DNA Sequencing The order of nucleotides in a DNA sequence is determined 1970s: performed manually using dye-terminator substances Now performed using dyes attached to nucleotides, with a laser and computerized machine to determine sequence 21.3 DNA Technology

7. 7 Automated DNA sequencer and an electropherogram Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Small section of a genome (sequencer): © Lawrence Berkeley Nat’l Lab/Roy Kaltschmidt, photographer Figure 21.14 Automated DNA sequencer and an electropherogram. 21.3 DNA Technology

8. 8 Polymerase chain reaction (PCR) Polymerase chain reaction is used to _______ small pieces of DNA. It is important for amplifying DNA for analysis such as in DNA fingerprinting. 21.3 DNA Technology

9. 9 Polymerase chain reaction (PCR) Figure 21.15 The polymerase chain reaction. 21.3 DNA Technology Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 3´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ DNA is denatured Into single strands 1. Sample is first heated to denature DNA. 2. DNA is cooled to a lower temperature to allow annealing of primers. 3. DNA is heated to 72°C, the optimal temperature for Taq DNA polymerase to extend primers. Cycle 2: 4 copies Cycle 3: 8 copies DNA strand DNA segment to be amplified Primers anneal to DNA Taq DNA polymerase 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 5´5´ 3´ 5´5´ 5´5´ 5´5´ 5´5´

10. 10 DNA fingerprinting Fragments are separated by their charge/size ratios Results in a _______ _______ for each individual Often used for paternity testing, or to identify an individual at a crime scene or unknown body remains Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Collect DNA Use gel electrophoresis to identify criminals markerSuspect BSuspect A Crime scene suspect B suspect A crime scene evidence Perform PCR on repeats 16 repeats12 repeats 16 repeats12 repeats Figure 21.16 PCR and electrophoresis used for DNA fingerprinting. 21.3 DNA Technology

11. 11 Gene cloning Recombinant DNA – contains DNA from 2 or more different sources that allows genes to be cloned Bacteria used to clone the human insulin gene –Restriction enzyme is used to cut the vector (plasmid) and the human DNA with the insulin gene. –DNA ligase seals together the insulin gene and the plasmid. –Bacterial cells take up plasmid, the gene is copied, and the product can be made. 21.3 DNA Technology

12. 12 Visualizing gene cloning Figure 21.17 Cloning of a human gene. 21.3 DNA Technology Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Bacteria produce a product. bacterium human DNA human cell insulin gene 4a4b DNA ligase seals the insulin gene into the plasmid. Gene cloning occurs. insulin 1 2 3 recombinant DNA plasmid (vector) Host cell takes up recombined plasmid. Restriction enzyme cleaves DNA.

13. 13 Biotechnology products: Transgenic organisms Important uses –Production of: Insulin Human growth hormone (HGH) Clotting factor VIII Tissue plasminogen activator (t-PA) Hepatitis B vaccine –Naturally-occurring oil-degrading bacteria can be made more effective through genetic engineering 21.3 DNA Technology

14. 14 Biotechnology products: Transgenic organisms Figure 21.18 Transgenic organisms. 21.3 DNA Technology a. b. c. d. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. a: © Maximilian Stock Ltd./Photo Researchers; b: Courtesy Robert H. Devlin, Fisheries and Oceans Canada; c: © Richard Shade; d: © Jerry Mason/Photo Researchers