MICRO-s10_13 - BIOL 240 General Microbiology General Spring...

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Unformatted text preview: 3/8/2010 BIOL 240: General Microbiology General Spring 2010 Rm. 22-116 T, Mar. 9, 2010 http://www.smccd.edu/accounts/staplesn/biol240/ 1. Pre-Lab Writeups: Be sure to prepare before each Lab W riteups Be Monday’s labs (for BOTH Mon. & Wed.)!! Monday – (What? Why? How? are we doing in the lab??) 2. Be sure to keep up with BY ARRANGEMENT HOURS!! Be They are REQUIRED for your grade!! • -- average TWO documented hours/week. 3. MT1 M/C Answer KEY is posted. Check and Check report any corrections by TODAY! report • under “Add’l Materials” tab. under 4. Bring DRY SOIL sample for your group of 6! Bring group 1. Compare transport and final electron acceptors between aerobic and two different types 1. Compare of anaerobic respiration. of 2. Explain how lipids and proteins are catabolized and energy harvested thru pathways Explain catabolized and shared with glucose metabolism. shared 3. Explain how light energy is harvested and stored iin the form of chemical energy by n Explain light photosynthetic organisms. photosynthetic 4. ** Distinguish the carbon and energy sources for all of the trophisms: chemo, photo, ** trophisms hetero, auto, and comb’ns of each. hetero, and comb 5. Diagram how catabolic and anabolic pathways can share intermediates to efficiently Diagram catabolic to regulate energy storage, energy usage, and biosynthesis…… regulate REVIEW: REVIEW: TODAY’s Objectives: Students should be able to... Students 1. Ch. 6: Describe several physical and chemical requirements for Describe microbial growth, and explain what factors determine optimal conditions. microbial 2. Define the terms that describe organisms that prefer high, medium, or low Define levels of each physical factor affecting growth. levels 3. Diagram and define the four phases of a bacterial growth curve. Diagram phases 4. Compare several methods of measuring microbial growth. Compare measuring 1 3/8/2010 5.12) Anabolism: Metabolic Metabolic Pathways of Energy Use Pathways A. Polysaccharide B. Lipid Biosynthesis Polysaccharide Biosynthesis Biosynthesis Figure 5.29 Figure 5.30 Anabolism/Biosynthesis C. Amino Acid and Protein Biosynthesis Figure 31 2 3/8/2010 Anabolism D. Purine and Pyrimidine Biosynthesis and Pyrimidine Figure 5.32 5.13) Amphibolic pathways • Are metabolic Are pathways that have both catabolic and catabolic and anabolic anabolic functions. Figure 5.33 3 3/8/2010 Chapter 6 Microbial Microbial Growth Growth Microbial Growth • Microbial growth = increase in increase number of cells, not cell size number 4 3/8/2010 6.1) Requirements for Growth: 6.1) Physical Requirements Physical A. Temperature: (Psychro-, meso-, thermo-philes) meso – – – Minimum growth temperature Optimum growth temperature Maximum growth temperature Figure 6.1 The Requirements for Growth: The Physical Requirements Physical B. pH: (acido-, neutero-, alkalo-philes) neutero alkalo – Most bacteria grow between pH 6.5 & 7.5. – Molds and yeasts grow between pH 5 & 6. – Acidophiles grow in acidic environments. Figure 6.4 C. Osmotic Pressure: – Hypertonic environments -- iincrease salt or sugar cause plasmolysis. Hypertonic -- ncrease cause plasmolysis – Extreme or obligate halophiles require high osmotic pressure. Extreme halophiles – Facultative halophiles tolerate high osmotic pressure . Facultative halophiles 5 3/8/2010 6.2) Requirements for Growth: 6.2) Chemical Requirements Chemical 1. Carbon – Structural organic Structural molecules, energy source molecules, – Chemoheterotrophs use use organic carbon sources organic – Autotrophs use CO2 3. Sulfur – In amino acids, thiamine, In biotin biotin – Most bacteria Most decompose proteins decompose – Some bacteria use SO42 or H2S – In DNA, RNA, ATP, and In membranes membranes – PO43 iis a source of s phosphorus phosphorus 2. Nitrogen – In amino acids, proteins – Most bacteria decompose Most proteins proteins – Some bacteria use NH4+ or NO3 – A few bacteria use N2 iin n nitrogen fixation nitrogen 4. Phosphorus 5. Trace Elements – Inorganic elements -- iin Inorganic -- n small amounts small – Usually as enzyme Usually cofactors cofactors The Requirements for Growth: The Chemical Requirements Chemical • Oxygen (O2) (Eg: BHIA deep agar tubes) 6 3/8/2010 Toxic Forms of Oxygen 1. Singlet oxygen: O2 boosted to a higher-energy energy state state 2. Superoxide free radicals: O2 • • er i s ut se O2 O2 2H supoxde dima H2O2 O2 3. Peroxide anion: O22 H+ 4. Hydroxyl radical ( OH) 6.3) Culture Media • Chemically Defined Media: Exact chemical Exact composition is known composition – “minimal media” – many additives for “fastidious” species many • Complex Media: Extracts and digests of Extracts yeasts, meat, or plants yeasts, – Nutrient broth – Nutrient agar 7 3/8/2010 A. Selective Media • Suppress unwanted Suppress microbes and encourage desired microbes. microbes. – EMB – MacConkey – NaCl-Mannitol – Min. Glc/Nitrate Min. Glc Figure 6.10 B. Differential Media • Make it easy to distinguish colonies of Make different microbes. different – [All the examples on previous slide] – Fermentation tubes – Blood agar, YE-CaCO3-sucrose, Snyder deep Figure 6.9 8 3/8/2010 6.4) Bact. Growth: Binary Fission 6.4) Bact. Figure 6.12 A. Cell Division: A. Exponential Increases!! Exponential 2n, where n = # cell division cycles Figure 6.13b 9 3/8/2010 B. Bacterial Growth Curve Figure 6.15 Question: What is happening to the cells at each phase?? C. Direct Measurements of C. Microbial Growth Microbial 1. Plate Counts: Perform serial Perform dilutions of a sample. dilutions – After incubation, After count colonies on plates that have 25-250 colonies 25 250 (CFUs) -- [we used -- [we 30-300 CFUs] 30 300 CFUs Figure 6.16 2. Cytometer count: Figure 6.20 10 3/8/2010 3. Most Probable Number Test Pos. 5 • Multiple tube MPN Multiple test test • Based on “dilution Based dilution to extinction” to – Highest dilution with Highest growth used to estimate bact. In liquid sample liquid 3 1 Figure 6.19 • Count positive tubes Count and compare to statistical MPN table. statistical D. Estimating Bacterial D. Numbers by Indirect Methods Numbers • Turbidity Figure 6.21 11 3/8/2010 Chapter 8 Microbial Genetics Terminology 1. Genetics: Study of what genes are, how they carry Study information, how information is expressed, and how genes are replicated; “the science of heredity” 2. Gene: Segment of DNA that encodes a functional Segment product, usually a protein product, 3. Genome = All of the genetic material in a cell 4. Genomics = Molecular study of genomes 5. Genotype = Specific forms of genes in an organism – Types of alleles present. 6. Phenotype = physical characteristics resulting from expression of the genes expression 12 3/8/2010 E. coli The Model Organism for molecular biology. Figure 8.1a Flow of Genetic Information: The Central Dogma DNA (m)RNA Protein Figure 8.2 DNA ** Central Dogma of Molecular Genetics ** ** Central 13 3/8/2010 DNA DNA 1. Polymer of nucleotides: Polymer 8.1) 8.1) adenine, thymine, cytosine, guanine (ATGC) cytosine, 2. Double helix associated Double with proteins with 3. "Backbone" is "Backbone" deoxyribose-phosphate deoxyribose 4. Strands held together by Strands hydrogen bonds between A=T and GC A=T and 5. Strands are Antiparallel Strands Antiparallel • 5’ (PO4) 3’ (OH) polarity Figure 8.3 DNA • Semi-Conservative Conservative Replication: Replication – Each “parental strand” Each serves as template… serves template – for synthesis of a new for “daughter strand” – Following complementary Following base-pairing rules base • A=T • G≡C – Rules allow one to predict Rules 2nd strand sequence from strand 1st!!! Figure 8.3 http://ncc.gmu.edu/dna/repanim.htm 14 3/8/2010 DNA Replication • 5’ 3’ synthesis • Template read Template 3’ 5’ Figure 8.5 DNA Replication 1. Open helix at Origin, lay-down primers: a) DNA Helicase “melts” strands apart, breaking H-bonds DNA Helicase b) Single-Strand Binding Proteins (SSB) keep template strands apart. keep c) RNA Primase llays down first several nucleotides (RNA!!) – gives “starting block” RNA Primase ays gives (free 3’-OH) to begin actual DNA synthesis. [Primers are removed later!] 2. DNA = copied by DNA polymerase III (Dpol3) DNA DNA 3. In the 5 3 direction – new nucleotides added to the 3’ In hydroxyl (-OH) group on deoxyribose in the growing strand OH) deoxyribose – • • Initiated by an RNA primer (RNA Primase enz.) Initiated RNA RNA Primase f ollows fork (1/fork; 2/ “bubble”) follows Opposite to fork movement Okazaki fragments (unsealed lagging pieces) 4. Leading strand synthesized continuously synthesized 5. Lagging strand synthesized discontinuously synthesized discontinuously 6. RNA primers are removed and Okazaki fragments joined by RNA DNA polymerase I & DNA ligase “fill and seal” to finish job!! DNA ligase http://bioteach.ubc.ca/TeachingResources/MolecularBiology/DNAReplication.swf http://www.wiley.com/college/pratt/0471393878/student/animations/dna_replication/index.html 15 3/8/2010 DNA Replication Fork Figure 8.6 III Leading Helicase Primase http://nobelprize.org/educati onal_games/medicine/dna/a/ replication/lagging_ani.html I Lagging http://www.stolaf.edu/people/giannini/flashanimat/molgenetics/dna-rna2.swf http://www.johnkyrk.com/DNAreplication.html http://www.wehi.edu.au/education/wehi-tv/dna/replication.html DNA replication is DNA Semiconservative & Bidirectional Semiconservative Bidirectional • Replication results in two Replication daughter DNA duplexes, •each with one each completely new completely strand, & strand, •one old strand strand (parental strand) (parental • = “SEMI-CONSERVATIVE” Figure 8.6 http://www.wehi.edu.au/education/wehi-tv/dna/replication.html • Two replication forks replication forks move in opposite move directions = •“Replication Bubble” 16 ...
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