Bio 425 Microbiology Dr. Ronald K. Sizemore Center for Marine Sciences 962-2304. Aimed at premed., predent., Grad. School - not for nurses Assumes solid Senior-level Biology background content heavy Good note taking skills needed Attendance as needed Seating Chart to help me try to learn names Interesting Lab where you will learn many basic Biology skills.
Bio 425 Microbiology Dr. Ronald K. Sizemore Center for Marine Sciences 962-2304. Aimed at premed., predent., Grad. School - not for nurses Assumes solid Senior-level Biology background content heavy Good note taking skills needed Attendance as needed Seating Chart to help me try to learn names Interesting Lab where you will learn many basic Biology skills.
Bio 425 Microbiology Dr. Ronald K. Sizemore Center for Marine Sciences 962-2304. Aimed at premed., predent., Grad. School - not for nurses Assumes solid Senior-level Biology background content heavy Good note taking skills needed Attendance as needed Seating Chart to help me try to learn names Interesting Lab where you will learn many basic Biology skills.
Center for Marine Sciences 962-2304 Sizemorer@uncw.edu How to catch me? - Email - Campus office - Friday 1002A
or
- Visit the Center!! Introduction to Bio 425 Aimed at Premed., Predent., Grad. School not for nurses Assumes solid Senior-level Biology background Content heavy Good note taking skills needed Attendance as needed Seating Chart to help me try to learn names Interesting Lab where you will learn many basic Biology skills My Goals and Expectations My Goals: To teach an organized, interesting course To give fair tests which are graded and returned promptly To cover the major content of Microbiology and share my personal experiences in Microbiology
My Expectations: I expect Senior-level work Attendance and read text as necessary Do not disrupt class (i.e. tardiness, leaving class early, etc) Misc. Text - Brock Biology of Microorganisms 11 th
Lab- Bensons Microbial Applications 11 th Short Edition - MUST be NEW or UNMARKED Test Dates dont change 1 Dropped Test allowed with prior approval of instructor Grade on 10 point scale no +/- grades Use my website for additional information http:/people.uncw.edu/sizemorer/rkswww/
Personal Information Name: email address: Major: Class: Relevant Classes: Career Plans Outside Influences: Microbiology Study of Germs? Bacteria - Bacteriology
Fungi Study of Fungi - Mycology Viruses Study of Viruses Virology
Algae
Study of Algae Phycology Immunology Study of the immune response Virus nonliving Bacteria living cells Size think small! Prokaryotes No Nucleus 2 Kingdoms - Domains Bacteria Archaea ?? Prokaryotes Eukaryotes
- Contain Nucleus
- Higher Organisms?
- More Complex Laboratory Culture Pure Culture Contamination Culture Media Petri Dishes Agar Sterilize Aseptic Technique Molecular Techniques Why Study Microbiology?
-Understand infectious Diseases Medicine -Understand Environmental Processes Biogeochemistry -Industrial Applications Biotechnology Bio 425 Word List Microbiology Archaea Culture Media Bacteriology Eukaryotes Petri Dishes Virology Nucleus Agar Immunology Viruses Sterilize Mycology Aseptic Phycology Laboratory Technique Germs Culture Diseases Living Cells Pure Biogeo- Prokaryotes Culture chemistry Bacteria Contamination Biotechnology Microscopes Compound Bright- Field Light Microscope Maximum Useful Magnification = 1200X Why? Resolution Useful magnification is limited by Resolution Resolution is influenced by Numerical Aperture (NA) NA determined by Wavelength of Light and Refractive Index Improve Magnification Shorten Wavelength Increase Refractive Index Staining Simple Stains Light Microscope - Colored Dye Electron Microscope - Electron Dense Differential Stains Gram Stain Gram Stain Most Important Stain Key Bacteriological Test Conventional Stain Multiple Steps & Subjective Fluorescent Stain One Step & +/- Optical Tricks Oil Immersion Lens
Dark Field Microscope Other Microscopes Phase Contrast Microscopes Fluorescent Microscopes Electron Microscopes Shorten wavelength Scanning Electron Microscope- SEM 3 Dimensional images TEM Better Resolution
Transmission Electron Microscope Microscopes Word List Compound Staining Confocal Light Simple Scanning Microscope Differential Laser Bright-Field Gram Stain Electron Magnification Phase Microscope Resolution Contrast Transmission Numerical Dark-field Scanning Aperture Fluorescent Oil Interference Immersion Contrast History Antoni van Leeuwenhoek Dutch Cloth Inspector 1684 wee animalcules Reported to the Royal Academy of Science - 1822- 1895 French Chemist Chance Favors the Prepared Mind - Spontaneous Generation 1864 Swan-necked Flask Ignored Spores
Louis Pasteur John Tyndall - Helped Pasteur Twice 1. Tyndall box Air 2. Tyndallization Method to sterilize heat sensitive materials Boil Incubate Boil Incubate Boil Sterile Germ theory Pasteur Wine disease
Lady Montagu Lady Montagu , 1718 Introduced variolation to England Edward Jenner 1798 Smallpox vaccination Ignaz Semmelweis, 1850 Asked Doctors to wash hands between patients Robert Lister ,1867 Introduced antiseptics to hospital Robert Koch Anthrax Kochs postulates
Kochs Postulates 1. Suspected pathogen always present in diseased animal 2. Isolated microorganism in pure culture 3. Re-inoculate healthy organism with pure culture and cause disease 4. Re-isolate the pathogen in pure culture Began the Concept of Guinea Pig? Lead to Discovery of the Cause of Tuberculosis Chicken Cholera Pasteur at his best? Attempted Kochs postulates Used old (attenuated) cultures Re-inoculates Chicken Protective Vaccine Pasteurs Chicken Cholera Experiment 1 st Experiment Healthy Chickens Old Culture Fresh Culture Live Chicken Dead Chickens 2 nd Experiment Attenuated Vaccinated Virulent Rabies Vaccine Charles Chamberlain Filterable Viruses Farmers Cure? Dried Spinal Tissue Attenuated Virus? Joseph Meister First Rabies Vaccine Antimicrobial Agents Paul Ehrlich Agent 606 Domagk - Prontosil Alexander Fleming 1929 - Penicillin Alexander Fleming Selman Waksman, 1942 Streptomycin Rutgers's Waksman Institute
Modern Molecular Microbiology Watson&Crick 1953
Kohler & Milstein 1979 - Monoclonal Antibodies George Kohler Cesar Milstein Boyer & Cohen 1975 - Recombinant DNA Woese & Fox Archaea 1977 George Fox Carl Woese Stanley Pruisner 1981 Prions Genomics Kary Mullis 1988 Polymerase Chain Reaction PCR DNA copying
Craig Ventor & Hamilton Smith 1995 Bacterial Genome Human Genome 2000? Ed Delong Marine Archaea Proteorhodopsin SAR Metagenomics Craig Ventor Microbiology Timeline History Word List Antonio van Semmelweis Attenuation Leeuwenhoek Lister 1867 Chamberlain 1684 Robert Koch Viruses Spontaneous Anthrax 1876 Alexander Generation Kochs Fleming 1864 Postulates 1929 Louis Pasteur Tuberculosis Waksman Germ Theory Chicken Watson Lady Montagu Cholera Crick 1953 Jenner 1798 Vaccine 1975 George Kohler & Cesar Milstein, 1979 Monoclonal Antibodies Stanley Cohen & Herbert Boyer Recombinant DNA Carl Woese & George Fox, 1977 Archaea
Stanley Pruisner, 1981 Prions Kary Mullins, 1988 PCR Craig Ventor & Hamilton Smith, 1995 Human Genome Project 2000 Word list 2 Cell Biology Prokaryotic Cell Small ~ 1-2m 01-50 m Max.? No Nucleus Little Internal Structure Some are Motile Unicellular or Clusters Always Exceptions? E. fishelsoni - 600m Cell Shapes Bacillus Coccus Spirillum Spirochetes Appendaged Cell Clusters Pairs Diplo Diplococcus Chains Strepto.. Streptococcus Others Staphylococcus
Cytoplasmic Membrane Phospholipid Bilayer No Sterols Ester linkages Archaea Ether linkage Lipid monolayer Thermophiles Membrane Function Selective Permeability Transport Proteins Simple Transport
Group Translocation
Glucose PEP
Glucose 6 PO4 Pyruvate Transport linked to Glycolysis ABC System Transport Periplamsic-binding protein Membrane-spanning protein Kinase
Bacterial Cell Wall Peptidoglycan, Murein Layer n-acetylglucosamine n-acetylmuramic acid Peptide Cross Links Diaminopimelic acid d-Alanine 1-4 Linkage Gram Positive Cell Wall Multi-Track layer Teichoic Acid Lysozyme Protoplast Autolysins Archaea Pseudopeptidoglycan Gram Negative Envelope Outer Membrane Lipopolysaccharides Peptidoglycan Periplasmic space Cytoplasmic Membrane Gram Negative Outer Membrane Lipopolysaccharides Endotoxins Pyrogens Lipid A Core Polysaccharide O Polysaccharide Porins Periplasmic Space Prokaryotic DNA Closed Circles E. coli chromosome has 4.6 million Bases Supercoiled Haploid Sometimes Plasmids Flagella Polar flagella Lophotrichous Peritrichous flagella Flagellin Self Assembly
Cell Biology Word List Prokaryotic 0.1-50m Transport Cell Shape 600m Max? Proteins Bacillus Cytoplasmic Group Coccus Membrane Translocation Spirillum Phospholipid Cell Wall Spirochete Bilayer Peptidoglycan Appendaged Sterols N-acetyl- Strepto, Diplo, Archaea glucosamine Staphylo. Ether-links N-acetyl- Size? Permeability muramic acid Cell Biology Word List 2 Peptide Archaea Core Poly- Cross-links Pseudo- saccharide Diamino- peptidoglycan Porins pimelic acid Gram - Periplasmic D-Alanine Lipopoly- Space 1-4 linkage saccharide Autolysin Gram+ Outer Prokaryotic Teichoic acid Membrane DNA Lysozyme Endotoxin Closed Circles Protoplast Lipid A 4.6X10 6 Bases Cell Biology Word List 3 Supercoiled Phototaxis Acid Haploid Magnetotaxis Glycogen Plasmids Pili, Fimbriae Poly- Flagella Capsule, phosphate Polar Flagella Glycocalyx Metachromatic Peritrichous Slime layer granules Flagellin Inclusion Sulfur Self-Assembly Bodies Magnetosomes Chemotaxis Poly-- Gas Vesicles Tumbles-Runs hydroxybutyric Endospores Cell Biology Word List 4 Endospores Endosymbiotic Dipicolinic Theory Acid Eukaryotic Organelles Nucleus Mitochondria Chloroplasts Nutrition and Metabolism Metabolism combines: Anabolism Biosynthesis with Catabolism Energy Generation Linked by Coupled Reactions Metabolic Types Energy Generation: Trapping Light Energy Phototrophs Chemical Reactions Chemotrophs
Carbon Source: CO 2 Lithotrophs Organic Carbon Organotroph Chemoorganotrophs are also called Heterotrophs Chemolithotrophs Bacterial Nutrition Macronutrients: Carbon source Nitrogen source Phosphate Sulfur Potassium Magnesium Calcium Sodium Iron Some bacteria use Siderophores to procure iron Micronutrients: Trace elements Growth Factors? Culture Media Defined media known Composition Complex Media Organic mixture Agar to Solidify Enzymes Organic Catalysts Reduced Activation Energy How Enzymes Act Form an Enzyme- Substrate Complex
Active site of the Enzyme holds Substrate
Enzyme Structure Enzymes are Proteins with Active Sites Additional Non protein Groups: Cofactors Loosely Bound Cofactors - Coenzymes Tightly Attached Cofactors- Prosthetic Groups Oxidation Reduction Redox Reactions Oxidation Lose of Electrons Reduction Gaining Electrons Electron Donors Electron Acceptors Reduction Potential
Electron Transfer Electron Tower Based on Reduction Potential Electron Carrier Primary Electron Donor (Food??) Final Electron Acceptor (Waste??) Energy Conservation High Energy Compounds (Energy Carriers??) ATP Adenosine Triphosphate Coenzyme A Phosphoenolpyruvate - PEP
Energy Generation Substrate-level Phosphorylation Fermentation Respiration Oxidative Phosphorylation Electron Transport (Cascade) Lactic Acid Fermentation Catabolic Pathways Glycolysis Embden-Meyerhof Pathway 3 Stages 1. Rearrangement 2. Oxidation 3. Reduction PEP Fermentation
Occurs in the absence of O 2
Less ATP produced Many fermentation products Substrate acts as electron donor and acceptor
Electron Cascade Coenzyme NADH NADH Dehydrogenase Flavoproteins Quinones Cytochromes Catalase, Oxidase Oxidative Phosphorylation Proton Motive Force Metabolic Poisons Uncouplers Dinitrophenol Inhibitors Carbon monoxide Cyanide Carbon Flow Citric Acid Cycle Krebs Cycle TCA Cycle Tricarboxylic Acid Cycle Carbon Oxidation CO 2 Production Reduced Coenzymes Anaerobic Respiration Nitrate Sulfate Carbonate Iron Final Electron Acceptor other than Oxygen
Chemolithotrophy Hydrogen Sulfide Hydrogen Gas Iron Ammonia
Phototrophy Catabolism Metabolism Word List Metabolism Catabolism Anabolism Coupled Reactions Nutrition Phototroph Chemotroph
Micronutrients Trace Elements Growth Factors? Culture Media Defined vs. Complex Media Metabolism Word List 2 Enzymes Organic Catalysts Activation Energy Enzyme- substrate Complex Active Site
Prosthetic groups Coenzymes Oxidation-Reduction Redox Reactions Electron Donors Electron Acceptors
Reduction Potential Electron Tower Electron Carriers Primary Electron Donor Final Electron Acceptor Metabolism Word List 3 High Energy Compounds Adenosine Triphosphate - ATP Coenzyme A Fermentation Substrate-level Phosphorylation Respiration Oxidative Phosphorylation Electron Transport Glycolysis Embden-Meyerhof Pathway Stage I, II, III Rearrangement Oxidation Reduction
Metabolism Word List 4 Aerobic Respiration Electron Cascade NADH dehydrogenase Flavoproteins Quinones
Cytochromes Catalase Oxidase Proton Motive Force Oxidative Phosphorylation Inhibitors
Uncouplers Carbon Flow - Citric Acid Cycle aka TCA, Tricarboxylic, Krebs Carbon Oxidation CO 2 Production Metabolism Word List 5 Reduced Coenzymes Biosynthesis ? Anaerobic Respiration nitrate, sulfate, carbonate ferric iron
Microbial Growth Binary Fission Growth Rate Generation Time E. coli can double every 20 minutes Many Bacteria have 1-3 hr Generation Times Growth Curve Lag Phase Log Phase (Exponential) Stationary Phase Death Phase Cryptic Growth Measuring Bacterial Growth
Total Cell Counts Direct Microscopic Counts Petroff-Hauser Counting Chambers Viable Counts Plate Counts Spread Plate Sample Spread on Solid Agar Plates
Pour Plate Dilute Sample Suspended into Melted Agar Dilution Extinction Dilution MPN Most Probable Number Turbidity Measurement Spectrophotometer Culture Type
Temperature Minimum Optimal Maximum Temperature Ranges Cold Temperatures Psychrophile Psychrotolerant Freezing Preserves Bacterial Cultures Other Factors pH Low pH Used to Preserve Foods Acidophiles Alkaliphiles Buffers Used in Many Bacteriological Media Water Availability Drying Limits Bacterial Growth Halophiles Osmophiles Xerophiles
Oxygen
Aerobe Anaerobes Facultative Microaerophile Growth Word List Bacterial Growth Binary fission Growth rate Generation Time Exponential Growth Batch Culture Growth Curve Lag Phase, Exponential (log) Phase, Stationary Phase, Death Phase, Cryptic Growth
Measuring Growth Total Cell Counts Direct Microscopic Counts Petroff-Hausser Counting Chambers
Growth Word List 2 Viable Count Plate Counts Spread Plate Pour Plate Dilution Extinction Dilution MPN Most Probable Number Turbidity Measurements Continuous Culture Chemostat Controlling Growth Temperature Minimum, Optimal, Maximum Psychrophile (psychrotolerant) Mesophile, Thermophile
Growth Word List 3 Hyperthermophile Freezing pH Acidophile Alkaliphile Buffers Water Activity
Drug Resistance Conjugative R Plasmids Drug Misuse
Search For New Drugs New Analogs Designer Drugs Growth Control Word List Growth Inhibition Sterilization Bacteriostatic Bactericidal Disinfection Heat Sterilization Decimal Reduction Time Spores Autoclave 121 C, 15 lb, 15 Minutes Pasteurization 63-66 C for 30 Minutes 71 C for 15 Seconds Radiation Filter Sterilization Depth Filters Membrane (Millipore) Filters Nucleopore Filter
Growth Control Word List 2 Antimicrobial Agent Bactericidal, Fungicidal, Viricidal Selective Toxicity Minimum Inhibitory Conc. Tube Dilution Agar Diffusion Method
Growth Factor Analogs Sulfa Drugs Sulfanilamide Folic Acid Synthesis Quinolones Antibiotics Broad-Spectrum Antibiotic Growth Control Word List 3 Lactam Ring Penicillins Cephalosporins Aminoglycosides Streptomycin, Neomycin, Kanamycin, Gentamicin Macrolides Erythromycin Tetracyclines Antiviral Agents Interferons Nucleos(t)ide analogs Protease Inhibitors Fungal Control Drug Resistance Conjugative R Plasmids Drug Misuse
Growth Control Word List 4 Search for New Drugs New Analogs Designer Drugs Molecular Biology Eukaryotes vs. Prokaryotes Eukaryotes Nucleus Linear Chromosomes Introns Repeated Sequences Monocistronic mRNA-Polyproteins
Eukaryotes Exon Splicing Prokaryotes No Nucleus Circular Chromosome No Introns No Repeats? Polycistronic mRNA Prokaryotes Polycistronic mRNA Gene A Gene B Sequence A Sequence B DNA Polycistronic mRNA Protein A Protein B DNA Structure Double Helix Complimentary Bases Hydrogen Bonds Double Stranded Antiparallel Bent DNA Inverted Repeats Stem-Loops Hairpins Sticky Ends Melting DNA DNA Melting Temperature determined by the number of GC pairs vs. A=T pairs
Higher G+C content of the DNA the higher temperature needed to denature it
Hybridization involves melting DNA and then allowing DNA strands to Renature (reforms base pairs) DNA Hybridization DNA in Cells is Supercoiled and in Eukaryotes covered with Histones
Chromosome Painting Topisomerase breaks then reseals DNA strands Allows DNA unwinding Plasmids Plasmid are nonessential extrachromosomal genetic elements found mainly in bacteria Plasmid replicate independently of the host chromosome and are often used in Genetic Engineering Transposable Elements Transposable elements move within the genome Insertion Sequences- Small Contain only genes to allow them to move Transposons - Movable genes Transposable Viruses Restriction Endonuclease Restriction Endonuclease Attack Specific Palindrome Sequences Modification Systems Change Palindrome Regions to Protect Them
DNA Fingerprint Because of Restriction Enzymes Specificity in Cutting DNA, It Can be Used to Create Maps Bacterial DNA Replication Relies on Complementary Base Pairs
Semiconservative uses a Template DNA Replication Several Different DNA Polymerases are used All the DNA Polymerases Needs a Double Stranded Primers to Initiate DNA Synthesis DNA Replication DNA Synthesis Begins at Origin Creates 2 Replication Fork Moves Bi-directional to Create Theta Structure Initiation of DNA Synthesis DNA Synthesis Begins at the Origin with Unwinding of the DNA by Helicases DNA Strands Held Apart by Single- strand Binding Proteins
Primase (RNA Polymerase) Creates Double Strand Template to Permit DNA Polymerase to Begin DNA Synthesis
Later a Second DNA polymerase removes RNA piece and Insert DNA piece
Ligase fills gap leaving double strand DNA Discontinuous DNA Replication Because DNA Replication Is Bi-directional and DNA Strands are Antiparallel: The Leading Strand Of DNA Replicates in a Continuous Manner The Other Lagging Strands Replication is Discontinuous and Produces Okazaki Fragments.
Leading & Lagging Strands Fragments Finally Glued Together by Ligase New DNA is Proofreads by Polymerases Transcription Transcription Produces RNA Copy of DNA Sequence RNA Polymerase Called Transcriptase Types of RNA Produced: mRNA Messenger RNA tRNA Transfer RNA rRNA Ribosomal RNA Beginning Transcription Promoter Pribnow Box (-10) Consensus Sequences (-10 & -35)
Sigma Factors Core Enzyme RNA Polymerase Termination of Transcription Rho Protein Stem-loop Nonbacterial Transcription 3 Eukaryotic RNA Polymerases Bacteria and Archaea have 1 RNA Polymerase But Archaea RNA Polymerase Most Closely Resembles Eukaryotic Polymerase Operon Operator + Structural Genes Polycistronic mRNA Bacterial Transcription RNA Processing Exons Introns Spliceosome Have Proteins Ribozyme Self Splicing Introns Primary Transcript Mature RNA Genetic Code Genetic Code Traits Codon 3 Bases Coding for Amino Acid Anticodon Base Pairs with Codon Code is Degenerate (more than one codon per amino acid) tRNA Wobbles on Codon Reading Frame
Stop Codons Start Codon AUG Set Reading Frame Shine-Dalgarno Region Open Reading Frame Translation
tRNA Charging tRNA Ribosomes 70S = 30S + 50S S = Svedberg units 30S = 16SRNA + 21 Proteins 50S = 5SRNA + 23SRNA + 34 Proteins Initiation of Translation Translate mRNA Sequence into Amino Acid Sequence Ribosome Attachment Shine-Dalgarno Region Start Codon n-Formylmethionine Process of Translation
Acceptor Site Peptide Site Elongation Termination of Translation Polysomes Termination Stop codon Ribosome Release Molecular Chaperone Translation: Alternatives Signal Sequences Posttranslational Modification Antibiotics Overlapping Genes Universal Code
Molecular Biology Eukaryotes vs DNA Structure Antiparallel Melting DNA Prokaryotes Double Helix Major/Minor GC, A=T Pairs Nucleus ? Complimentary Grove Hybridization Linear-Circular Bases Bent DNA Supercoiled Chromosome Hydrogen Inverted Loops Histones Intron? Bonds Stem-Loop Topoisomerase Histones? Double Hairpins Genetic Repeats Stranded Sticky Ends Elements Molecular Biology 2 Plasmids Endonucleases DNA Primer? Transposons Palindrome Replication Initiation of Insertion Modification Complementary DNA Synthesis Sequences Restriction Base Pairing Origin Viruses Enzyme Semi- Replication Restriction/ (Fragment) conservative Fork Modification Map Template Bi-directional Restriction DNA Polymerases Theta Structure Helicases Molecular Biology 3 Single-strand Ligase rRNA = Consensus Binding Proteins Proofreading Ribosomal RNA Sequence Primase = RNA RNA Termination Polymerase Transcription Polymerase Rho Proteins Leading Strand mRNA = Promoter Archaea? Continuous Message RNA Sigma Factors Polycistronic Discontinuous tRNA = Core Enzyme mRNA Okazaki Fragments Transfer RNA Pribnow Box Operon Molecular Biology 4 Operon Genetic Code Start codon = Acceptor Site Operator Codon AUG Peptide Site Anticodon Reading Frame Polysomes RNA Processing Degeneracy Shine-Dalgarno Termination Splicesome Wobble Region Stop Codon Ribozyme Stop Codon n-formyl- Molecular Exons Introns Nonsense methionine Chaperones Codon Elongation Signal Sequence Molecular Biology 5 Postranslational Modifications Antibiotics Overlapping Genes Universal Code Gen Expression Constitutive Enzymes not Regulated Most Enzymes Can Have Their Activity Regulated Feedback Inhibition Allosteric Sites Isozymes Covalent Modification Regulation of Transcription Operator Operon
Control Negative Control Positive Control Activator Protein Regulon DNA Binding Proteins Histones Attenuation Attenuator Leader Sequence Catabolite Repression Global control Diauxic growth Catabolite Repressor Protein cAMP
Other Global Controls Quorum Sensing Alternative Sigma Factors Proteases Chaperonin Two Component Regulation
Signal Transduction Sensor Kinase Response Regulator Others Chemotaxis Antisense RNA Triple DNA Regulation of Gene Expression Constitutive Isozymes Corepressor Enzymes Covalent Operator Regulation of Modification Operon Enzyme Regulation of Repression Activity Transcription Repressor Feedback Enzyme Protein Inhibition Induction Negative Control Allosteric Site Inducer Operator Gene Expression 2 Negative DNA Binding Catabolite Control Proteins Repression Positive Attenuation Catabolite Control Leader Activator Activator Sequence Protein Protein Attenuator cAMP Regulon Global Control Quorum Histones Diauxic Growth Sensing Gene Expression 3 Other Global Signal Antisense Controls Transduction mRNA Alternative 2-Component Triple DNA Sigma Factors Regulation Proteases Sensor Kinases Chaperonin Response Regulator Chemotaxis Microbial Genetics Mutation Genetic Recombination Microbial Models Molecular Cloning Biotechnology Golden Rice Mutants
Mutation Wild Type Genotype Phenotype Isolation of Mutants
Replica Plating Auxotrophs Nutritional Mutants Prototroph Wild Type
Penicillin Selection Method Kills Growing Cells Mutations Spontaneous Mutations Point Mutations Base-Pair Substitution Silent, Missense, or Nonsense Mutations Conditional Lethal Mutations Temperature Sensitive Mutations 30 C 40 C Incubation Temperature Reading Frame Frame Shift Mutation Back Mutation Suppressor Mutation Mutation
Prototroph Auxotroph
Back Mutation Mutation Back Mutation Normal Protein Other Mutations Deletions Lose of a Piece of DNA Insertions Additions of a Piece of DNA Translocation Movement of a Piece of DNA to a New Location in the Genome Inversion Orientation of a Piece of DNA is Reversed in the Genome
Mutation Rate Spontaneous (Natural or Background) Mutation Rate DNA ~ 1 per Million (varies between genes and location) RNA ~ 1 per Thousand Mutagens Anything that Increases the Mutation Rate Mutagens Chemicals Base Analogs Chemically Resemble Base
Intercalating Agent Inserts Between Base Pairs Radiation UV Light Ionizing Radiation DNA Repair SOS Regulatory System Biological Mutagens Transposons Can Insert Into Gene Site Directed Mutations Man-made Ames Test Mutagens Carcinogens Ames Test Measures Back Mutation Rates to Detect Carcinogens Bacterial Gene Transfer Genetic Recombination 3 Types of Bacterial Gene Exchange: 1. Transformation Naked DNA Competent
Transfection in Eukaryotes Particle (Gene) Guns Insert DNA 2. Transduction Generalized Transduction Specialized Transduction Lysogeny Can Lead to Phage Conversion 3. Conjugation
Plasmids Copy Number Incompatibility Curing Conjugative Plasmid Plasmid Types Resistance Plasmids R Plasmids Virulence Plasmids Bacteriocins Engineered Plasmids
Chromosomal Mobilization Fertility (F) Plasmid Episome HFr Strain Sex Pili Gene Maps Interrupted Matings Transposition Transposase Conservative Transposition Replicative Transposition
Phase Variation Inversion
E. coli Chromosome 4,639,221 Base Pairs 4,288 Open Reading Frames Horizontal Gene Transfer Pathogenicity Islands Genomics Mycoplasma genitalium Genome 470 Open Reading Frame Genomic Mining Genetic Word List Mutation Genetic Recombination Microbial Models Molecular Cloning Biotechnology Mutants
Genotype Phenotypes Wild-Type Isolation Mutants Replica plating Auxotroph Prototroph Penicillin-Selection Method Spontaneous Mutations Point Mutation Base-Pair Substitution Silent, Misssense, Nonsense Mutations Genetic Word List 2 Temperature Sensitive Mutation Reading Frame Frame Shift Mutation Back Mutations Suppressor Mutation
Mutagens Chemicals Base Analogs Intercalating Agents Radiation UV Light
Genetic Word List 3 Ionizing Radiation DNA Repair SOS Regulatory System Biological Mutagens Transposons Site Directed Mutations Carcinogensis Ames Test Genetic Recombination Bacterial Gene Transfer Transformation Competent Transfection Electroporation Particle gun
Genetic Word List 4 Transduction Generalized Transduction Specialized Transduction Lysogeny Phage conversion Conjugation Plasmids Copy Number Incompatibility Curing Conjugative Plasmids Resistance Plasmids R Plasmids Virulence Plasmids Bacteriocin Engineered plasmids
Genetic Word List 5 Chromosomal Mobilization Sex Pili Fertility (F) Plasmids Hfr Strains Interrupted Matings Genetic Maps
Transposition Transposase Conservative Replicative Transposition Phase Variation Inversion Escherichia coli Chromosome 4,639,221 Base Pairs 4,288 Open Reading Frames Horizontal Gene Transfer
Genetic Word List 6 Pathogenicity Islands Genomics Mycoplasma genitalium Genome 470 Open Reading Frames Genomic Mining
Viruses Individual Viruses = Virion Bacterial Viruses = Bacteriophage Genome = Double Strand or Single Strand DNA or RNA Human Papillomavirus Protein Coat Capsid, Coat, Shell Capsomer - Individual Proteins Self Assembly Nucleocapsid Viral Symmetry Icosahedral
Helical Icosahedral Subunits Tobacco Mosaic Virus Herpes Simplex Virus Enveloped Virus Complex Virus T 4 Bacteriophage Enzymes in Viruses Reverse Transcriptase Lysozyme RNA Polymerase Viral Host Living Cells Animal Host Cell Culture Tissue Culture Monolayer Cell Line Primary Culture Line Transformation Viruses Mutagen Permanent Cell line HELA
Quantification of Viruses Plaque Assay Bacteria Efficiency of Plating Animal Infectivity Methods End Point Dilution
Bacteriophage Plaques Plaques in Tissue Culture Monolayer Viral Replication
Attachment
Penetration Early Step - Produce Early Proteins Replication of Nucleic Acids Production of Late Proteins Assembly Release One-step Growth Curve Latent Period Some Host Cells Use Restriction Enzymes to Destroy Viruses Some Viruses Can Modify Their Genomes Restriction Sites for Protection +/- SS RNA Viruses SS DNA Viruses DS Intermediates Reverse Transcriptase Bacterial Viruses Bacteriophage Model Viruses Genetic Engineering +SS RNA Bacteriophage MS2 Male Specific Produces 4 Proteins Maturation Protein,
Coat Protein Lysis RNA Polymerase Overlapping Genes SS DNA Bacteriophage X 174 First Genome Sequenced in 1977 Overlapping Genes Replicative Form - Double Stranded DS DNA Bacteriophage Lytic T Phages T4, T7 Head, Tail, Spikes Base Plate, Tail Fibers, Contractile Sheath Hydroxymethyl- cytosine Lambda Best Known Virus? Can be Virulent or Temperate Lysis or Lysogeny Becomes Prophage (Provirus) Gives Immunity to Host Maintenance of Lysogeny Lambda Repressor CRO Protein Induction Transposable Phage Insertion Elements Transposon Mu Phage Mutator Phage Animal Viruses DNA Viruses RNA Viruses Eukaryotic Host Types of Infections: Lytic Persistent Latent Hepatitis A Herpes Cancer Viruses Transformation Neoplasm Benign
Papovavirus SV 40 Human papillomavirus Smallpox Adenovirus Retrovirus
RNA Genome Reverse Transcriptase DNA Copy Insert into Host Genome AIDS Acquired Immunodeficiency Syndrome HIV Human Immunodeficiency Virus HIV Viroids Naked Circles SS RNA 246-375 Nucleotides Plant Diseases
Prions Infectious Proteins? Chaperone Protein Kuru Mad Cow Disease Scrapies Creutzfeldt- Jakob Disease Viruses Word List Virion Bacteriophage 28 200 nm (0.2 m) Genome SS/DS RNA/DNA Capsid, Coat, Shell Capsomer Self-Assembly Nucleocapsid Viral Symmetry Helical Icosahedral Enveloped Viruses Complex Viruses Enzymes in Virion Reverse Transcriptase
Lysozymes Viral Host Living Organisms Cell (Tissue) Culture Monolayer Primary Culture Line Viruses 2 Transformation Permanent Cell Line Quantification of Viruses Plaque Assay Efficiency of Plating
Animal Infectivity Methods End Point Dilution Viral Reproduction Attachment Penetration
Early Steps (Early Proteins) Replication of Nucleic Acids Synthesis of Proteins (Late) Assembly Release Viruses 3 One-step Growth Curve Latent Period Restriction/ Modification Restriction Enzymes Modification +/- SS RNA Viruses
SS DNA Viruses DS Intermediates Reverse Transcriptase
BACTERIAL VIRUSES Model Viruses
Genetic Engineering RNA Bacteriophage +SS RNA Viruses MS2 Male Specific Phage
T Cell Receptors Haptens Antigenic Determinants Epitopes Nonspecific Immunity Phagocytes
Specific Immunity B Lymphocytes Antiserum Immunoglobulin
Gamma Globulin IgG Heavy Chains Light Chains Antigen Binding
Constant Region Variable Region Gene Rearrangement 2 Antigen Binding Sites per IgG Immunoglobulins IgG - Smallest, Most Common, Transplacental IgA - Secretory IgM - Largest IgD IgE - Allergies IgA IgM T Cells
T Lymphocytes T Cell Receptors Major Histocompatibility Complex Cytokines
Cytotoxic T Cells Natural Killer Cells Clonal Selection Immune Tolerance 1 st , 2 nd Response
Carriers Mortality Morbidity Disease Reservoirs Acute Carriers Incubation Period Chronic Carriers Transmission Host-to-Host Vector Fomites Food and Water Common-Source Epidemic John Snow Stopped Cholera Epidemic in London (1855) by Removing Broad Street Pump Handle Herd Immunity Nosocomial Infections Public Health Control Reservoir Prevent Transmission Immunization Quarantine Surveillance Pathogen Eradication Global Health Travel Developing Countries Reemerging Diseases Emerging Diseases
Epidemiology Word List Prevalence Incidence Endemic Epidemic Pandemic Carrier Mortality Morbidity Disease Reservoirs Acute Carrier Incubation Period Chronic Carrier Typhoid Mary Transmission Host-to-Host Vector
Fomites Food and Water Common-Source Epidemic John Snow - Cholera Herd immunity Nosocomial Infections Public Health
Epidemiology Word List 2 Control Reservoir Prevent Transmission Immunization Quarantine Surveillance Pathogen Eradication
Global Health Travel Developing Countries Reemerging Diseases Emerging Disease Person-to-Person Microbial Diseases Airborne Transmission Respiratory Infections Streptococcus pyogenes S. pneumoniae Strept Throat Other Strept Infections Hemolytic Strept. Otitis media Impetigo Scarlet Fever Rheumatic Fever Pneumonia Staphylococcus aureus Coagulase Positive Boils Impetigo Toxic Shock Syndrome
Other STDs Chlamydia trachomatis Lymphogranuloma venereum Herpes simplex I & II
AIDS Acquired Immunodeficiency Syndrome HIV Human Immunodeficiency Virus CD4
Reverse Transcriptase Protease Inhibitor
Progression of HIV P to P Diseases Word List Airborne Transmission Respiratory Infections Streptococcus pyogenes S. pneumoniae Strept Throat Hemolysis Otitis media
Impetigo Scarlet Fever Rheumatic Fever Pneumonia Staphylococcus aureus Coagulase positive Boils Impetigo Toxic Shock Syndrome Diphtheria Corynebacterium diphtheriae Metachromatic Granules P to P Diseases Word List 2 Pseudomembrane Whooping Cough Bordetella pertussis DPT Vaccine Tuberculosis Mycobacterium tuberculosis Primary Tuberculin Test Postprimary Tubercles M. bovis Hansens Disease M. leprae Leprosy Viral Respiratory Pathogens Common Cold Rhinovirus Influenza Hemagglutinin Neuraminidase P to P Diseases Word List 3 Antigenic Shift Polyvalent Vaccine Measles (Red) Rubeola Mumps MMR Vaccine German Measles Rubella Chickenpox Varicella Shingles Sexually Transmitted Diseases Gonorrhea
Neisseria gonorrheae Syphilis Treponema pallidum Congenital Syphilis Chancre Secondary Syphilis Tertiary Syphilis P to P Diseases Word List 4 Chlamydia trachomatis Lymphogranuloma venereum Herpes simplex I & II
Acquired Immunodeficiency Syndrome Human Immunodeficiency Virus CD4 Reverse Transcriptase Protease Inhibitors Transmitted Microbial Diseases Zoonoses Rabies Hantavirus Four Corners Disease Typhus Rickettsia prowazekii Other Rickettsial Diseases Rocky Mountain Spotted Fever R. rickettsii Ehrlichiosis Ehrlichia Q Fever Coxiella burnetii Lyme Disease Borrelia burgdorferi Deer Ticks Plague Bubonic plague Yersinia pestis Pnuemonic plague Rat Fleas Endemic (Sylvatic) Plague Common Source Diseases Water Quality Coliform Counts Extinction Dilution Most Probable Numbers MPN Fecal Coliform Membrane Filter Sewage Treatment
Primary Treatment Settling Tank Sludge Septic Tank Secondary Treatments Activated Sludge Trickling Filters Other Treatments
Biodisc Sludge Digesters Methane Tertiary treatment BOD
Biotechnology Molecular Cloning Recombinant DNA HISTORY Herb Boyer EcoR1 Stanley Cohen Competent Cells Plasmids Stanley Falkow Asilomar Conference 1975 Voluntary Band of Some Recombinant Research NIH-RAC formed as a result Isolate Gene Fragmented Total DNA
Use mRNA and Make DNA with Reverse Transcriptase Splice into Cloning Vehicle Plasmids Viruses Use Restriction Endonuclease to create Sticking Ends Rejoin Molecule with Ligase Inserting into Host
Transformation Detection of Successful Clone DNA Library Shotgun Cloning Cloning Vector Plasmid Cloning Vectors Single Cleavage Site High copy Numbers Insertion Inactivation Virus Vectors
Lambda Phage Vector Stable Packaging Cosmids Other Vectors Expression Vectors Secretion Vectors Shuttle Vectors Bacteriophage M13 DNA Sequencing Yeast Artificial Chromosome Human Genome Project Hosts
Cloning Vector Hosts E. coli Endotoxins Yeast Detection of Clones
Antibodies Nucleic Acid Probes Synthetic DNA Production of Recombinant Protein Transcription of Cloned Gene Promoter
Translation of Cloned Gene Ribosome Binding Site Polymerase Chain Reaction
Kary Mullins PCR DNA Amplification TAQ Enzyme Primer PCR Site Directed Mutagenesis Knockout Mutation Applications of Genetic Engineering
Human Insulin
Interferon
Hepatitis Vaccine Vaccines Subunit Vaccines Hepatitis Vaccine Recombinant Vaccines Polyvalent Vaccines DNA Vaccines Edible Vaccines Other Products Tissue Plasminogen Activator Rennin Ti Plasmid Bacillus thuringiensis BT Roundup Resistant Favosavr Tomato More Possibilities Ice Minus Bacteria PCB Degrading Bacteria Luminescent Reporter Gene Transgenic Animal Human Gene Therapy DNA Chips - Microarrays Actual DNA Chip Genetic Eng. Word List Biotechnology Molecular Cloning Recombinant DNA Isolate Gene mRNA, Reverse Transcriptase Splice into Cloning Vehicle - Plasmids, Viruses Restriction Endonucleases Sticky Ends Ligase Insert into Host Detect Successful Clone DNA Library Shotgun Cloning Plasmid Cloning Vectors Single Cleavage Sites
Genetic Eng. Word List 2 High Copy numbers Insertion Inactivation Lambda Phage Vector Stable Packaging Cosmids Other Vectors Expression Vectors Secretion Vectors Shuttle Vectors Bacteriophage M 13 DNA Sequencing Yeast Artificial Chromosomes Human Genome Project Cloning Vector Hosts E. coli Yeast Detection of Clones Antibody
Genetic Eng. Word List 3 Nucleic Acid Probes Transcription of Cloned Gene Promoter Translation of Cloned Gene Ribosome Binding Site Synthetic DNA Probes
Polymerase chain Reaction PCR DNA Amplification TAQ Enzyme Primers Site Directed Mutagenesis Knockout Mutations
Applications of Genetic Engineering Human Insulin Interferon Hepatitis Vaccine Recombinant Vaccine Subunit Vaccines
Genetic Eng. Word List 4 Polyvalent Vaccines DNA Vaccines Tissue Plasminogen Activator Rennin Ti Plasmid Bacillus thuringiensis Roundup Resistance Favosavr Tomato Ice Minus Bacteria PCB Degrading Bacteria Luminescent Reporter Gene Transgenic Animals
Aerobic Sparger Cooling Impellor Scale-up Antibiotics Antibiotic Production Search for New Antibiotics Screening Protein Engineering Natural, Biosynthetic, Semisynthetic Penicillins
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