MCB2004 Exam 2 Lecture 9 Microbial Growth Increase in the number of cells o 2 2 Generations etc o Exponential growth Bacterial Growth Curve o Lag phase Preparing for growth but no increase in population o Log phase exponential increase in population o Stationary deaths production of cells o Death population is decreasing at a logarithmic rate Physical Requirement temperature o Minimum optimum maximum growth temperature Physical requirement pH o pH log H o Optimum pH between 6 5 and 7 5 o Molds and yeast between 5 and 6 o Acidophiles acidic environments Physical requirement osmotic pressure o Hypertonic cause plasmolysis b c insides of cell leave cell wall Physical requirements Irradiation o Visible light between ultraviolet and infared Chemical requirements Energy for chemoheterotrophs and autotrophs Amino acids proteins Amino acids thiamine biotin DNA RNA ATP and membranes Found in small amounts enzyme cofactors o Carbon o Nitrogen o Sulfur o Phosphorus o Trace elements o Trophisms Energy Chemotrophs chemical Phototrophs light Carbon Autotrophs only use inorganic carbon Heterotrophs use organic compound s o Toxic forms of Oxygen reactive oxygen species Signlet O2 boosted to higher energy Superoxide free radicals O2 Peroxide anion O2 2 Hydroxyl radical OH Chemical requirements organic growth factors o Obtained from enviornemnt o Vitamins AA purines pyrimidines Culture media o Culture medium nutrients prepared for microbial growth o Sterile no living microbes o Inoculum introduction of microbes into medium o Culture microbes growing on medium o Chemically defined media exact chem Composition known o Complex media extracts of meat yeast or plant o Complex polysaccharide o Solidifying agent for culture media in petri o Not metabolized by microbes o Differential media distinguish colonies of different microbes Agar liquid medium plate Pure culture streak plate o One species or strain o Colony cells arising from single cell CFU o of bacteria mL CFU x reciprocal of dilution sample Viable count Direct count MPN Turbidity o of cells x 1250000 o Count positive tubes and match with chart o Light shining through tube to dector Lecture 10 Control of microbial growth Humans control growth of microbes o Decrease to o Increase to Ferment food Make medicine Recycle Biological control Prevent spoilage disease contamination Important terminology o Sterilization removal of all forms of microbial life o Commercial sterilization heat treatment to kill endospores o Disinfection destruction of vegetative pathogens o Antisepsis destruction on living tissue o Degerming removal of microbes from limited area o Sanitation lower microbial count Death kinetics o Bacteria populations die at a constant rate Physical methods of microbial control o heat Denatures proteins Autoclave steam under pressure Pressure increases as temp increases o Pasteurization Reduce spoilage High temp for a short time Thermoduric organisms survive o Dry heat sterilization Kills by oxidation o Filtration o Radiation Removes microbes Damages DNA Concentration Organic matter pH Time Disinfectants Ionizing radiation nonionizing radiation o Chemical methods of microbial control Nitrate prevent endospore germination Antibiotics nisin prevent spoilage of cheese Biguanides surgical hand washing and skin prep Alcohols denature proteins dissolve lipids Heavy metals Sterilants Aldehydes inactivate proteins Gaseous denature proteins o Resistance of microbes Most resistant Prions Least resistant virus with lipid envelopes Lecture 11 Microbial genetics Terminology o Genetics what genes are how replicated info is expressed o Gene DNA segement o Genome all genetic material in a cell o Genomics study of genomes o Genotype genes of an organism o Phenotype expression of genes Father of genetics o Mendel o Pea plant experiments Gene expression o Replication Transcription translation DNA o AT and GC o Backbone deoxyribosephosphate o Held together by hydrogen bonds o Semiconservative replication o Copied by DNA polymerase o Lagging strand Okazaki fragments o PCR Joined by polymerase and ligase Multiple copies of one piece of DNA Cloning amplify DNA diagnose disease detect pathogens RNA o Transcription Starts when RNA polymerase binds to promoter Stops when reaches terminator sequence 5 3 direction o Translation mRNA protein Translated into codons Start AUG Stop UAA UAG UGA 64 codons encode 20 AA Genetic code is degenerate tRNA carries anticodon Operon o PO control region promoter operon o If repressor binds to operon transcription will be turned off Mutation Change in genetic material o Terminology Mutagen agent that causes mutations Spontaneous happens in absence of mutagen o Types Base substitution Change in one base Missence Nonsense Change in AA Makes stop codon Frameshift Insert or delete at least one pair of nucleotides o Ames test Incubation of rat liver extract with mutagen Test for carcinogens Conjugation o F F 2F o HFr F no change Plasmids o Conjugative plasmid carries gene for sex pili and transfer of plasmid o Dissimilation of plasmidsL encode enzymes for catabolism o R factors encode antibiotic resistance Transposons o DNA that can move from one region to another o Cut and reseal DNA o Can carry other genes Lecture 12 Biotechnology and Recombinant DNA Recombinant DNA technology insertion or modification of genes to produce desired Biotechnology use of microorganisms cells or cell components to make a product Advantages of microbial organisms proteins o Grow quickly o Well understood Restriction enzymes o Cut specific places on DNA o Destroy harmful bacteriophages o Cannot digest host DNA because of methylates cytosines o RM restriction modification system primitive defense mechanism Vectors o Carry new DNA to desired cells o Plasmids viruses can be used as vectors Reverse transcriptase digests mRNA takes out introns Proton expression and purification o Put protein coding sequence in plasmid clone o Transform plasmid into bacteria o Grow bacteria induce promoter o Lyse cells Taxol Plant product used for treatment for ovarian cancer produced in E Coli Transfer foreign DNA into cells o Transformation Bacteria o Transfection mammalian cells o Electroporation o Microinjection o Gene gun o Viral vectors Viral gene therapy o Replace the gene necessary for pathogenic growth List at least five applications of rDNA technology o Crop protection o Viral gene therapy cure disease permanently o Human insulin o Immunize against pathogens o Replace
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