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Chapter 5: DNA, Gene Expression, and Biotechnology What is the code and how is it harnessed? Lectures by Mark Manteuffel, St. Louis Community College Learning Objectives Describe what DNA is and what it does. Explain the process of gene expression and the collaboration of nature and nurture. Explain the causes and effects of damage to the genetic code. Describe biotechnology and its implications for human health. Discuss biotechnology today and tomorrow. Discuss biotechnology in agriculture. 5.1 “The DNA 200” Knowledge about DNA is increasing justice in the world. What is the most common reason why DNA analyses overturn incorrect criminal convictions? Take-home message 5.1 DNA is a molecule that all living organisms carry in every cell in their body. Unique in virtually every person, DNA can serve as an individual identifier, left behind us as we go about our lives. This is a fact that is used increasingly to ensure greater justice in our society, such as through establishing the innocence of individuals wrongly convicted of crimes. Two Important Features of DNA (1) DNA contains the instructions on how to create a body and control its growth and development. (2) The instructions encoded in the DNA molecule are passed down from parent to offspring. Rosalind Franklin DNA “Double Helix” – a nucleic acid DNA consists of individual units called nucleotides: a sugar, a phosphate group, and a nitrogencontaining base. How does knowing the structure of DNA help us? • Manipulating DNA allows us to manipulate genetic traits (proteins) and treat genetic disorders (gene therapy). • HIV treatment • Biomimicry Helix Wind Turbine – quieter, smaller, less impact on birds and bats – residential scale. AZT mimics the chemical shape of Thymine, and when HIV viral DNA tries to replicate, AZT attaches to block. • The genetic code is shared by all organisms. • This tobacco plant is expressing a firefly gene. • The gene codes for the firefly enzyme (a protein) that causes it to glow. The number of chromosomes varies from species to species. Corn has 10 unique chromosomes. Fruit flies have only 4. Dogs have 39 chromosomes. Goldfish have 47 chromosomes. Humans have 46 chromosomes. Individuals in each of these species inherit one copy of each chromosome from each parent. 5.3 Genes are sections of DNA that contain instructions for making proteins. DNA is a universal language that provides the instructions for building all the structures of all living organisms. Alleles are alternate versions of a gene that code for a the same trait. Usually they occur in pairs as dominant and recessive alleles. 5.4 Not all DNA contains instructions for making proteins. Why doesn’t that make an amoeba more complex than a human? The Proportion of the DNA That Codes for Genes: Just because we don’t yet know it’s function doesn’t mean non-coding DNA has no function. Introns = Noncoding regions of DNA May take the form of short (or long) sequences that are repeated thousands of times, may also consist of gene fragments or duplicate versions of genes. FROM EGG TO ORGANISM: HOW AND WHY GENES ARE REGULATED • Four of the many different types of human cells – They all share the same genome – What makes them different? (a) Three muscle cells (partial) (b) A nerve cell (partial) (c) Sperm cells (d) Blood cells The flow of information from genes to proteins (genotype to phenotype) is called gene expression. In the mature organisms, each cell type has a different pattern of turned-on genes. Genotype = all of the genes contained in an organism Phenotype = the physical manifestations of the instructions 5.6 Transcription: reading the information coded in DNA In transcription, a single copy of one specific gene within the DNA is made, in the form of a molecule of mRNA, which moves where it can be translated into a protein. 5.7 Translation: using information from DNA to build usable molecules In translation, the information from a gene that has been carried by the nucleotide sequence of an mRNA is read, and ingredients present in the cell’s cytoplasm are used to produce a protein. Several ingredients must be present in the cytoplasm for translation to occur. Transfer RNA Free amino acids Ribosomal units The amino acids of a polypeptide correspond with the nucleotides of a mRNA molecule. This is how translation occurs. Amino acids are organized by triplets called codons. Overview: – Transcription occurs in the nucleus. – Translation occurs in the cytoplasm. • A mutation – Any change in the nucleotide sequence of DNA can have a wide range of effects. Mutant hemoglobin DNA mRNA mRNA Normal hemoglobin Glu Sickle-cell hemoglobin Val Figure 10.21 Breast Cancer in Humans Two human genes, called BRCA1 and BRCA2 More than 200 different changes in the DNA sequences of these genes have been detected, each of which results in an increased risk of developing breast cancer. Types of Mutations – Can be divided into two general categories • Base substitution • Nucleotide deletion or insertion – Can result in positive, negative, or nonfunctional changes in proteins. Met Lys Phe Gly Ala Met Lys Phe Ser Ala Figure 10.22a Mutations plan an important role in evolution. Take-home message 5.9 Most genetic diseases result from individual mutations that cause a gene to produce a non-functioning enzyme, which in turn blocks the functioning of a metabolic pathway. A “fastflush” response • Mutations may result from – Errors in DNA replication (spontaneous mutations) – Physical or chemical agents (mutagens) VIRUSES: GENES IN PACKAGES • Viruses sit on the fence between life and nonlife. – They exhibit some but not all characteristics of living organisms – They cannot reproduce outside of a host cell. Figure 10.24 A virus uses the host cell’s “machinery” to transcribe and translate its own DNA or RNA; it takes over the host cell, multiplies, bursts, and releases new viral particles into the host organism. 5.10 What is biotechnology? Genetic Engineering Adding, deleting, or transplanting genes from one organism to another, to alter the organisms in useful ways This creates recombinant organisms or genetically modified organisms. How is Genetic Engineering different from Artificial Selection? • Natural Selection – Requires environmental pressures, genetic variability, and successful reproduction. • Artificial Selection – breeding plants and animals for desired traits • Genetic Modification/Engineering – Creating new genetic combinations (recombinant organisms) in the lab – Crossing kingdom (and species) boundaries Overview: Cut out the gene of interest, paste it into a bacterial plasmid, and test the colony for gene expression. Step 1: Use Restriction Enzymes to recognize a specific sequence and cut out the gene of interest. Different enzymes are used to cut at different sequences. Step 2: Use PCR to amplify or clone the gene of interest. The DNA polymerase enzyme is used instead of the RNA polymerase because we’re making more DNA instead of transcribing mRNA, but the process is similar. Step 3: Paste the gene of interest into bacteria DNA (a circular plasmid). Incorporate the plasmid into the bacteria cell using a gene gun, a viral promoter, or electrical shock. The DNA ligase enzyme pastes the ends together. Cell containing gene of interest Host cell 1 DNA Recombinant DNA (host DNA plus gene of interest) 2 Gene of interest Cell multiplies and produces protein 3 Genes may be inserted into another organism Protein may be harvested OR Gene for pest resistance inserted into plants Protein dissolves blood clots in heart attack therapy Biotech advances in human health fall into three categories: (1) producing medicines to treat diseases (2) curing disease (3) preventing diseases from occurring in the first place. By transferring the gene for a desired protein product into a bacterium, proteins can be produced in large quantities. Curing diseases with biotechnology Using Stem Cells and Gene therapy and the correction of malfunctioning genes Stem Cells: Embryonic and Adult • Embryonic stem cells are undifferentiated cells in the early animal embryo that give rise to specialized cells. Grown in the laboratory, certain growth factors can induce changes in gene expression so that the cells may develop into a certain cell type. • Adult stem cells are partially differentiated cells present in adult tissues in order to generate replacements for specialized cells and tissues. – More difficult to grow in laboratory culture – Can be harvested from adult patients avoiding the controversy surrounding the embryo and with less risk of immune system rejection. Therapeutic Cloning and Stem Cells HUMAN GENE THERAPY • Human gene therapy is a recombinant DNA procedure that seeks to treat disease by replacing the mutant version with a properly functioning one. Why has gene therapy had such a poor record of success in curing diseases? (1) Difficulty getting the working gene into the specific cells where it is needed. (2) Difficulty getting the working gene into enough cells and at the right rate to have a physiological effect. (3) Problems with the transfer organism getting into unintended cells (virus side effects) Is a given set of parents likely to produce a baby with a genetic disease? Prevention has to do with screening parents, developing fetuses, growing children, and adults Ethical Dilemmas Discrimination Genetic 2008 Health Information Nondiscrimination Act of insurance Does not cover life, disability or long-term care insurance How to proceed with the information? Should the parents terminate the pregnancy? 5.17 DNA as an individual identifier: the uses and abuses of DNA fingerprinting What is a DNA fingerprint? • The goal of DNA fingerprinting by is to determine whether or not samples of DNA contain identical genetic markers. Genetic markers are unique unless you have an identical twin. Crime scene 1 DNA collected 2 DNA amplified if necessary 3 DNA cut into fragments 4 DNA fragments compared Suspect 1 Suspect 2 Figure 12.13 Crime scene DNA Suspect DNA Fragment w Cut Fragment z Fragment x Cut Fragment y Cut Fragment y Figure 12.16 – Gel electrophoresis is a method for sorting these fragments based on their length and electrical charge. Mixture of DNA fragments of different sizes Longer (slower) fragments Power source Gel Shorter (faster) fragments Completed gel Figure 12.17 • The DNA fragments are visualized as “bands” on the gel – The bands of different DNA samples can then be compared. Crime scene DNA Suspect DNA Longer fragments Shorter fragments Figure 12.18 • DNA fingerprints from a murder case • Guilty or innocent? • This blood on the defendant’s clothes matches the victim. Blood on defendant’s clothes Defendant’s blood Victim’s blood 5.18 DNA sequences reveal evolutionary relatedness (1) Mapping Genetic Landscapes: The Human Genome Project (2) Building Earth’s Family Tree When we say humans and chimps are genetically 96% identical, what do we mean? An indication that chimps are our closest living relative. Remember most of that DNA doesn’t code for functional proteins. When comparing only the coding parts of the genome, we find only 29% of our genes code for the same exact proteins found in chimps. 5.19 The promise and perils of cloning From organs (therapeutic cloning) to individuals (reproductive or whole organism cloning) • Differentiated plant cells – Have the ability to develop into a whole new organism (produce plant clones – reproductive cloning) Root of carrot plant Plantlet Cell division in culture Single cell Root cells in growth medium Adult plant Reproductive Cloning of Animals – Involves replacing nuclei of egg cells with nuclei from differentiated cells – Has been used to clone a variety of animals FDA has approved cloned animals for meat and milk. Dolly, March 1997 March, 2002 Nov., 2001 May, 2003 Health problems are associated with cloned species. These little guys did not live a full natural life. ANDi, Rhesus monkey GM’d to carry a fluorescent protein from a jellyfish Jan., 2001 Is it possible to clone a dinosaur? If so, how could it be done? 5.14 Producing more nutritious and better food with biotechnology How might a genetically modified plant help 500 million malnourished people? Nutrientrich “golden rice” Putting vaccines in potatoes… How can genetically modified plants lead to reduced pesticide use by farmers? But what about the pesticides in our diet? Herbicide Resistance Are plants like LibertyLink Corn and Roundup Ready Soy good for the ecosystem? Farmers still pay the costs. Faster Growth and Bigger Bodies Do we want more growth hormones in our diet? What happens if these giant fish escape? Almost everyone in the U.S. consumes genetically modified foods regularly without knowing it. Do we have a choice? Fears and risks: the safety of genetically modified foods? Fear #1. Organisms that we want to kill may become invincible. How long before the pests evolve to become resistant to the pesticide gene in plants? Fear #2. Organisms that we don’t want to kill may be killed inadvertently. Fear #3. Genetically modified crops are not tested or regulated adequately. Fear #4. Eating genetically modified foods is potentially dangerous. The long-term effects are unknown. Allergies may develop. Fear #5. Have we lost consumer freedom? If GM crops are not labeled and there is a health problem, it will be very hard to prove accountability. Fear #6. Loss of genetic diversity among crop plants is risky. Fear #7. Hidden costs may reduce the financial advantages of genetically modified crops. Fear #8. These combinations “matings” could never occur in nature. Fear #9. Recombinant DNA technology could create hazardous new pathogens (i.e. if cancer cell genes were transferred into an infectious bacteria or virus). Is it even ethical to patent living organisms?