Biology, 9th ed,Sylvia Mader Chapter 16 Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. DNA probe array tagged DNA did bind to probe DNA probe tagged DNA did not bind to probe tagged DNA testing subject's DNA

Biology, 9th ed,Sylvia Mader Chapter 16 Outline Biotechnology and Genomics DNA Cloning Recombinant DNA Technology Restriction Enzyme DNA Ligase Polymerase Chain Reaction Biotechnology Products Transgenic Bacteria, Plants, and Animals Gene Therapy Genomics

Biology, 9th ed,Sylvia Mader Chapter 16 DNA Cloning Biotechnology and Genomics Cloning is the production of identical copies of DNA Members of a bacterial colony on a petri dish are clones because they all came from division of the same cell. Identical twins are clones Single embryo separate to become two. Gene cloning is production of many identical copies of the same gene. If the inserted gene is replicated and expressed, we can recover the cloned gene or protein product. Cloned genes have many research purposes Humans can be treated with gene therapy

Recombinant DNA Technology Biology, 9th ed,Sylvia Mader Chapter 16 Recombinant DNA Technology Biotechnology and Genomics Recombinant DNA (rDNA) contains DNA from two or more different sources Requires: A vector introduces rDNA into host cell Plasmids (small accessory rings of DNA from bacteria) are common vectors Two enzymes are required to introduce foreign DNA into vector DNA A restriction enzyme - cleaves DNA, and A DNA ligase enzyme - seals DNA into an opening created by the restriction enzyme

Biology, 9th ed,Sylvia Mader Chapter 16 Cloning a Human Gene Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. human DNA plasmid bacterium human cell O f r o m a i 1. Restriction enzyme cleaves DNA. insulin gene 2. DNA ligase seals human gene and plasmid. recombinant DNA 3. Host cell takes up recombined plasmid. 4. Gene cloning occurs. insulin

Biology, 9th ed,Sylvia Mader Chapter 16 Restriction Enzyme Biotechnology and Genomics Cuts DNA at specific points. Cleaves vector (plasmid) and foreign (human) DNA. Cleaving DNA makes DNA fragments ending in short single-stranded segments with “sticky ends.” The “sticky ends” allow insertion of foreign DNA into vector DNA. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. A G A A T T C G C DNA duplex T C T T A A G C G restriction enzyme A A T T C G C "sticky ends" G C G A G T C T T A A

Biology, 9th ed,Sylvia Mader Chapter 16 DNA Ligase Biotechnology and Genomics Seals the foreign gene into the vector DNA Treated cells (bacteria) take up plasmids Bacteria and plasmids reproduce. Many copies of the plasmid and many copies of the foreign gene.

DNA Cloning: Polymerase Chain Reaction (PCR) Biology, 9th ed,Sylvia Mader Chapter 16 DNA Cloning: Polymerase Chain Reaction (PCR) Biotechnology and Genomics Amplifies a targeted sequence of DNA Create millions of copies of a single gene or a specific piece of DNA in a test tube Requires: DNA polymerase Withstands the temperature necessary to separate double-stranded DNA. A supply of nucleotides for the new, complementary strand

Biology, 9th ed,Sylvia Mader Chapter 16 PCR Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. PCR cycles DNA copies first 1 DNA double strand second 2 new strand old strand third 4 new old fourth 8 old new fifth 16 and so forth

Applications of PCR: Analyzing DNA Segments Biology, 9th ed,Sylvia Mader Chapter 16 Applications of PCR: Analyzing DNA Segments Biotechnology and Genomics DNA fingerprinting is the technique of using DNA fragment lengths Treat DNA segment with restriction enzymes A unique collection of different fragments is produced Gel electrophoresis separates the fragments according to their charge/size Produces distinctive banding pattern Usually used to measure number of repeats of short sequences Used in paternity suits, rape cases, corpse ID, etc.

DNA Fingerprinting & Paternity Biology, 9th ed,Sylvia Mader Chapter 16 DNA Fingerprinting & Paternity Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Mother Child Male 1 Male 2 few Base repeat units many a. DNA Band patterns Fluorescence units Increasing size b. Automated DNA fingerprinting

Biotechnology Products Biology, 9th ed,Sylvia Mader Chapter 16 Biotechnology Products Biotechnology and Genomics Genetically engineered organisms can produce biotechnology products. Organisms that have had a foreign gene inserted into them are transgenic.

Biology, 9th ed,Sylvia Mader Chapter 16 Transgenic Bacteria Biotechnology and Genomics Gene of interest is inserted into bacteria. Bacteria are grown in large vats called bioreactors and product is harvested. Products on the market include insulin, hepatitis B vaccine, t-PA, and human growth hormone. Transgenic bacteria can produce chemical products. Transgenic bacteria process minerals. Metals Collection

Biology, 9th ed,Sylvia Mader Chapter 16 Transgenic Bacteria Biotechnology and Genomics Transgenic bacteria promote plant health Bacteria that colonize corn roots can be endowed with genes for insect toxin Transgenic bacteria can degrade substances. Oil-Eating Bacteria

Genetically Engineered Bacteria Biology, 9th ed,Sylvia Mader Chapter 16 Genetically Engineered Bacteria Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. (Both): Courtesy General Electric Research & Development

Biology, 9th ed,Sylvia Mader Chapter 16 Transgenic Plants Biotechnology and Genomics Agricultural Crops Foreign genes now give cotton, corn, and potato strains the ability to produce an insect toxin Soybeans are now resistant to a common herbicide Human Hormones Plants are being engineered to produce human proteins including hormones, clotting factors, and antibodies in their seeds

Biology, 9th ed,Sylvia Mader Chapter 16 Transgenic Animals Biotechnology and Genomics Vortex Mixing: Many types of animal eggs have taken up the gene for bovine growth hormone (bGH) The procedure has been used to produce larger fishes, cows, pigs, rabbits, and sheep Gene Pharming: Use of transgenic farm animals to produce pharmaceuticals Genes coding for therapeutic & diagnostic proteins are incorporated into an animal’s DNA The proteins appear in the animal’s milk Plans are to produce drugs to treat Cystic fibrosis Cancer Blood diseases, etc.

Biology, 9th ed,Sylvia Mader Chapter 16 Transgenic Mammals Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. human gene for growth hormone microinjection of human gene donor of egg development within a host goat human growth hormone Transgenic goat produces human growth hormone. milk a. transgenic goat cells with gene for human growth hormone microinjection of transgenic gene into enucleated donor eggs enucleated eggs donor of eggs development within host goats Cloned transgenic goats produce human growth hormone. milk b.

Biology, 9th ed,Sylvia Mader Chapter 16 Transgenic Animals Biotechnology and Genomics Researchers are using transgenic mice for various research projects. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. one-celled mouse embryos with two X chromosomes no injection inject SRY DNA Embryo develops into a female. Embryo develops into a male. FEMALE MALE

Biology, 9th ed,Sylvia Mader Chapter 16 Gene Therapy Biotechnology and Genomics Gene therapy involves procedures to give patients healthy genes to make up for a faulty gene. It also includes the use of genes to treat genetic disorders and various human illnesses. There are ex vivo (outside body) and in vivo (inside body) methods of gene therapy. Ex Vivo Children with Severe Combined Immunodeficiency Bone Marrow Stem Cells In Vivo Cystic Fibrosis Nasal / Respiratory Spray

Biology, 9th ed,Sylvia Mader Chapter 16 Gene Therapy Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Brain (gene transfer by injection)* • Huntington disease • Alzheimer disease • Parkinson disease • brain tumors Skin (gene transfer by modified blood cells)** • skin cancer Lungs (gene transfer by aerosol spray)* • cystic fibrosis • hereditary emphysema Liver (gene transfer by modified implants)** • familial hypercholesterolemia Blood (gene transfer by bone marrow transplant)* • sickle-cell disease Endothelium (blood vessel lining) (gene transfer by implantation of modified implants)** • hemophilia • diabetes mellitus Muscle (gene transfer by injection)* • Duchenne muscular dystrophy Bone marrow (gene transfer by implantation of modified stem cells)** • SCID • sickle-cell disease * in vivo ** ex vivo

Biology, 9th ed,Sylvia Mader Chapter 16 Genomics Biotechnology and Genomics Genomics is the study of genomes of humans and other organisms. Sequencing the Bases The Human Genome Project produced a working draft of all the base pairs in all chromosomes. Took 13 years to sequence three billion base pairs along the length of chromosomes.

Biology, 9th ed,Sylvia Mader Chapter 16 Human Genome Project Biotechnology and Genomics Genome - All the genetic information of an individual (or species) Goals of Human Genome Project Determine the base pair sequence Construct a map showing sequence of genes on specific chromosomes

Biology, 9th ed,Sylvia Mader Chapter 16 Human Genome Project Biotechnology and Genomics Humans have 20,500 genes Most code for proteins Much of the human genome was formerly described as “junk” Does not specify the order of amino acids in a polypeptide Recent observation suggest that between 74% and 93% of the genome is transcribed into RNA Thus, vast junk DNA wasteland may be much more important than once thought

Eukaryotic Gene Structure Biology, 9th ed,Sylvia Mader Chapter 16 Eukaryotic Gene Structure Biotechnology and Genomics Historically, genes were defined as discrete units of heredity that corresponded to a locus on a chromosome. Prokaryotes typically possess a single circular chromosome. Eukaryotic chromosomes are much more complex.

Biology, 9th ed,Sylvia Mader Chapter 16 Chromosomal DNA Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. intergenic sequences Gene C exon exon DNA intron Gene A Gene B exon exon pre-mRNA intron RNA introns 5¢ 3¢ 5¢ 3¢ 3¢ 5¢ Gene A mRNA Gene B mRNA Gene C mRNA © Cindy Charles/PhotoEdit

Eukaryotic Gene Structure Biology, 9th ed,Sylvia Mader Chapter 16 Eukaryotic Gene Structure Biotechnology and Genomics An Intergenic Sequences are DNA sequences that occur between genes Repetitive DNA elements occur when the same sequence of two or more nucleotides are repeated many times along the length of one or more chromosomes. Transposons are specific DNA sequences that have the remarkable ability to move within and between chromosomes.

Genomic Differences Between Species Biology, 9th ed,Sylvia Mader Chapter 16 Genomic Differences Between Species Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

Biology, 9th ed,Sylvia Mader Chapter 16 HapMap Project Biotechnology and Genomics People inherit patterns of sequence differences, called haplotypes If one haplotype of a person has an A rather than a G at a particular location in a chromosome, there are probably other particular base differences near the A Genetic data from African, Asian, and European populations will be analyzed A HapMap is a catalog common sequence differences that occur in a species The goal of the project is to link haplotypes to risk for specific illnesses May lead to new methods of preventing, diagnosing, and treating disease

Functional and Comparative Genomics Biology, 9th ed,Sylvia Mader Chapter 16 Functional and Comparative Genomics Biotechnology and Genomics Functional genomics aims to understand the role of genome in cells or organisms DNA microarrays can monitor the expression of thousands of genes simultaneously and tell us: What genes are turned on Environmental conditions that turn on the gene DNA microarrays contain microscopic amounts of known DNA fixed onto a small glass slide mRNA bind through complementary base pairing mRNA is produced by active cell. Identify various mutations in the genome of an individual This is called the person’s genetic profile.

DNA Microarray Technology Biology, 9th ed,Sylvia Mader Chapter 16 DNA Microarray Technology Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. DNA probe array tagged DNA did bind to probe DNA probe tagged DNA did not bind to probe tagged DNA testing subject's DNA

Biology, 9th ed,Sylvia Mader Chapter 16 Genetic Profile Biotechnology and Genomics The complete genotype of an individual This is the person’s genetic profile A way of studying how genes work together to control the phenotype Analyze the genetic profile of many individuals Compare their profiles to their phenotypes

Biology, 9th ed,Sylvia Mader Chapter 16 Proteomics Biotechnology and Genomics The study of the structure, function, and interaction of cellular proteins At least 25,000 of our genes are translated into proteins The sum total of these proteins is called the human proteome Understanding protein function is essential to the development of better drugs Correlate drug treatment to the particular genome Increase efficiency and decrease side effects Once the primary structure of these protein is known It should be possible to predict their tertiary structure Computer modeling of the tertiary of these proteins is an important part of proteomics

Biology, 9th ed,Sylvia Mader Chapter 16 Bioinformatics Biotechnology and Genomics The application of computer technologies to the study of the genome Genomics and proteomics produce raw data These fields depend on computer analysis to find significant patterns in the data Scientists hope to find relationships between genetic profiles and genetic disorders New computational tools will be needed to accomplish these goals

Biology, 9th ed,Sylvia Mader Chapter 16 Bioinformatics Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. intergenic sequences Gene C exon exon DNA intron Gene A Gene B exon exon pre-mRNA intron RNA introns 5¢ 3¢ 5¢ 3¢ 3¢ 5¢ Gene A mRNA Gene B mRNA Gene C mRNA

Biology, 9th ed,Sylvia Mader Chapter 16 Review Biotechnology and Genomics DNA Cloning Recombinant DNA Technology Restriction Enzyme DNA Ligase Polymerase Chain Reaction Biotechnology Products Gene Therapy Genomics

Biology, 9th ed,Sylvia Mader Chapter 16 Biotechnology and Genomics Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. DNA probe array tagged DNA did bind to probe DNA probe tagged DNA did not bind to probe tagged DNA testing subject's DNA