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358 Cards in this Set
- Front
- Back
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Ubiquitous
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Can be found just about everywhere.
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aerotolerance
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ability or inability to live in the presence of oxygen
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aerobic metabolism byproducts
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H2O and CO2
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Anaerobic metabolism byproducts
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Nitrate,
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Where do Obligate aerobes grow in broth tube
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At the top
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Facultative anaerobes
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can live with or without oxygen. Grow in middle of the broth
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Obligate anaerobes
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Do not require oxygen, inhabit lower regions of broth
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Capnophile
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Can survive only if CO2 levels are elevated
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Thioglycollate broth
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has indicature resazurin.
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Resazurin
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Indicator of oxygen presence. Turns pink in presence of oxygen.
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Aerobic, anaerobic, or facultative?
P.aeruginosa S. aureus C. sporogenes |
P. aeruginosa= aerobic
S. aureus= facultative C. sporogenes= anaerobic |
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Fluid Thioglycollate
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Liquid medium to promote growth of a wide variety of microorganisms. associated with anaerobic bacteria/ prevent rapid infusion of . O2
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Purpose of Anaerobic Jar
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To create an oxygen free environment
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What activates the gas pack?
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10 cc of water
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what does the gas pack contain?
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sodium borohydride and sodium bicarb, methylene blue paper strip to indicate presence of oxygen
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What is the catalyst included in the GasPak?
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Palladium. produce the necessary conditions. Catalyzes rx between hydrogen and free oxygen to produce water.
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What color is methylene blue if O2 is present? Absent?
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Blue = Present
Colorless = absent |
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What are some reducing agents?
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Fluid thioglycollate medium, ascorbic acid, and cysteine
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What macroelements or macronutrients do bacteria require? Which in larger amounts than in humans?
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S, P, O, N, C, H Ca, Mg, Fe
Mg, Fe |
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microelements (trace elements)
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Mn, Zn, Co, Mo, Ni, Cu
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what trace elements are used by bacteria as cofactors or enzymes?
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Ni, Cu
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How are trace elements supplied?
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By water or in media components.
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Lithotroph
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Electrons from reduced inorganic material
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What macroelements or macronutrients do bacteria require? Which in larger amounts than in humans?
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S, P, O, N, C, H Ca, Mg, Fe
Mg, Fe |
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microelements (trace elements)
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Mn, Zn, Co, Mo, Ni, Cu
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what trace elements are used by bacteria as cofactors or enzymes?
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Ni, Cu
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How are trace elements supplied?
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By water or in media components.
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Lithotroph
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Reduced get sources of carbon, Energy and Electrons from reduced inorganic material
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Organotroph
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Electron source is organic molecules. Majority of Bacteria are organotrophic
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Autotroph
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Use CO2 as principle carbon source. Must obtain energy from other sources
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Heterotroph
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Use organic molecules as carbon and energy source
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Phototroph
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Use light as energy source
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Chemotroph
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Use oxidation of organic or inorganic compounds as energy source.
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Sources of Nitrogen
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Organic molecules
Ammonia Nitrate via assimilatory nitrate reduction Nitrogen gas via N fixation |
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Primary example of assimilatory nitrate reduction bacteria
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E. Coli
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Sources of Phosphorus and Sulfer
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Phosporus- Inorganic phos. incorporated into cells direct.
Sulfer- Sulfate reduction by assililitory sulfate reduction |
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Most nutrients enter the cell by:
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Facilitated diffusion
Active transport Group translocation |
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Major way bacteria pick up nutrients
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Active transport
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Passive Diffusion
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Molecules move from higher conc. to lower. rate dep. on size and concentration gradient. Not used much by bacteria
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What moves across via passive diffusion?
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H2O, O2, and CO2
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Facilitated diffusion
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Similar to Passive, NOT energy dependent. Is is enzyme mediated. Not used much by bacteria. Uses permeases.
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What is effectively transported by facilitated diffusion?
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Glycerol, sugars, amino acids
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Major way substance brought into bacteria cell
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Group Translocation
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2nd major way transport molecules into cell
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Standard active transport
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Best known system of Group translocation
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Phosphoenolpyruvate: sugar
Phosphotransferase System (PTS) |
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Siderophores
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Aid in iron uptake
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Why do some bacteria need high iron concentrations?
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To produce ferradoxin used as an electron carrier
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Where is ferradoxin usually found?
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Plants- major electron carrier in photosynthetic reactions
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ferraquats
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bind feradoxin by interrupting photosynthesis. Involved in uptake of of Iron siderophores. Needed in substrate level, not cofactor level.
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General purpose media example
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Nutrient broth
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Enriched media example
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Blood and blood products
Brain/Brain tissue |
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Selective media
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Allows for growth of certain organisms
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What Selective medium will ONLY Staphlococci grow?
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High Salt medium
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What is defined media?
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Media where all components and concentrations are known
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Complex media
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some ingredients of unknown composition and/or concentration.
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What is carbon source in complex medua
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lactose
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Crystal violet prevents what from growing on media?
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Gram Positive organisms
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What complex agar is differential and selective?
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MacConkey Agar
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Types of complex media
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Nutrient broth
Tryptic Soy broth MacConkey agar |
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Peptones
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Protein hydrolysates prepared by partial digestion of various protein sources. Differ dep. on protien
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what is agar made from
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Seaweed
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Where do extracts come from for media components
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Beef or yeast
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Supportive or General purpose media
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Support growth of many microorganisms. Eg. TSA
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What media is supplemented by blood or other special nutrients?
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Enriched media Eg. Blood agar
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Chocolate agar
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hemolyzed RBC's.
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What microrganism requires chocolate agar to grow
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Moraxella catarrhalis
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What color is Moraxella catarrhalis on chocolate agar?
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White
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alpha hemalysis
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Breaking down of RBC's releasing Hgb
Greenish darkening under colonies |
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Beta hemolysis
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Break down or RBC and Hgb.
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Types of selective and differential media
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Blood agar
Eosin methylene blue (EMB)agar MacConkey agar Mannitol Salt agar |
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nutrient agar and salt
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Selective media that only grows staphlococci
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What does mannitol do and what does it define?
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It ferments the sugar and allows differentiation between S. aureus and S.epidermis.
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In mannitol agar, what is the pH indicator, and what color is it in prsence of acidic contents?
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pH indicator is phenol red.
turns yellow if acidic |
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In mannitol, what color is medium with staph aureus? Why?
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It is yellow because of the release of acidic products.
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Differential media allows you to distinguish groups base on what?
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Biological characteristics
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blood agar is what type of media?
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Enriched differential medium
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MacConkey agar is what type of medium?
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Selective differential
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What does blood agar differentiate between?
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hemolytic vs nonhemolytic bacteria
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What does MacConkey agar differentiate?
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Lactose fermenters vs. nonfermenters
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Pure Culture
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Population of cells arising from a single cell
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Where does most rapid colony growth occur?
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On edge of colony
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What do bacteria need to do to form a biofilm?
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attach. Biofilms produced by most organisms
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How do most bacteria grow?
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Binary fission
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What growth do microbiologist usually study?
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Population growth rather than individual cell growth
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What 2 pathways function during Prokaryotic cell cycle?
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1.DNA replication and partition
2. cytokinesis |
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what type of DNA do bacteria have?
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Circular DNA
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what is the suspectad mechanism for segregation?
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Actin and tubulin filaments
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Mesosome
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Site of attachment of bacteria chromosome to cytoplasmic membrane.
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Origin of replication
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site where replication begins
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Terminus
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site replication terminated
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replisome
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group of proteins needed for DNA synthesis.
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cell division may be single, _________, __________, or _________
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tetrads, chains, or clusters
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protein MreB
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plays role in cell shape and movement of chromosomes to opposite poles
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protein FtsZ
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plays a role in Z ring formation essential for separation
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How long does cell cycle take?
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Completed in 20 minutes
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How long for DNA replication?
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40 minutes
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How long for septum formation and cytokinesis?
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20 min
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How can cell cycle be 20 min if seems to take 60 min.?
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Second, third, or fourth round begin before first round completed.
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Batch Culture and what Temp?
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culture incubated in closed vessel with a single batch of medium.
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Growth curve usually has how many planes? What are they?
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Four.
1. Lag 2.exponential (log) 3. stationary 4.Death |
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how is growth usually plotted?
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Logarithm of cell number vs time. NOT LINEAR!!!!!!!
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What does the Oxidase test detect
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bacteria that produce cytochrome c oxidase. cytochrome c oxidase is found in ETC (aerobic met.)
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chromogenic reagent used in oxidase test
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tetramethyl-p-phenylenediamine
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chromogenic reducing agents produce color if
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cytochrome c is present
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Oxidase test can be used to differentiate
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oxidase neg. Enterobacteriaceae from oxidase positive pseudomonadaceae
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dropps of reagent on oxidase positive bacteria produce what color?
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Purple-blue
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What is Nitrogen reduction test used to identify
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anaerobic, gram - bacteria that contain nitrate reductase
Ex enerobacteriaceae |
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denitrification
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multi-step process of creating nitrate to nitrite
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Nitrate reductase
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enzyme capaple of one step reduction of nitrate to nitrite
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What type of medium is Nitrate broth and components
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Undefined- beef extract, peptone, and potassium nitrate.
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Color reactions in nitrate broth are due to
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result of reactions btwn metabolic products and reagents added after incubation.
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What is evidence of denitrification?
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presence of gas in the Durham tume and organism is not a fermenter.
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if no evidence of denitrification, what should be done?
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Add sulfanilic acid and naphthylamine to test for reduction to nitrite. if present forms nitrous acid-red
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Simmons Citrate Agar
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medium that contains citrate as sole carbon source and ammonium phosphate as sole nitrogen source
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What is added a indicator in Simmons citrate agar
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Bromtymol blue dye- green at 6.9 and blue at 7.6
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What indicates a positive citrate test?
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change in color of the medium from green to blue.
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What is the color change caused by in citric acid test?
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conversion of ammonium phosphate to ammonia and ammonium hydroxide
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Growth with absent color change in citrate test is considered
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evidence of positive reaction
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Decarboxylation Test
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Used to differentiate members of enterobacteriaceae and distinguish them from other gram- rods
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Decarboxylases catalyze reactions that produce
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alkaline products
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What does Moller's Decarboxylase Base Medium contain
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peptone, glucose, and pH indicator Bromcresol purple and coenzyme pyridoxal phosphate
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What color is Bromcresol purple at pH 6.8? Below 5.2?
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6.8 and above- purple
5.2 and below- yellow |
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In Decarboxylation medium inoculation, why is mineral oil used to seal the medium
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To seal from external oxygen and promote fermentation.
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Glucose fermentation in decarboxylation anaerobic medium turns the medium
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Yellow because accumulation of acid endproducts.
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Decarboxylation of the amino acid results in end products that turn the medium
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Purple.
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What color does medium turn and stay in presence of glucose fermenter producing inappropiate decarboxylase?
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Yellow
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What color will medium be if organism does not ferment glucose in decarboxylation test
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There will be no color change
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what is the only positive result in decarboxylation test?
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Purple
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Decarboxylase media are used to differentiate organisms in the family _________ and distinguish them from
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Enterobacteriaceae
Gram negative rods |
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Products of a deamination reaction
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ammonia (NH3) and phenylpyruvic acid.
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What does the reagent added to phenylalanine agar after innoculation contain?
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Ferric Chloride
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Reaction of phenylpyruvic acid and ferric chloride turns what color?
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Dark Green.
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Formation of green color in phenylalanine deaminase test indicates presence of
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phenylpyruvic acid (also means phenylalanine deaminase is present)
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What color indicates a negative Phenylalanine deaminase test?
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Yellow
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Whay type of medium is Phenylalanine deaminase test medium?
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Defined,
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waht is phenylalanine deaminase test used to differentiate?
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Morganella, Proteus, and Providencia from other members of Enterobacteriaceae
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What type of medium is Bile Esculin Agar?
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Both selective and differential. It is also undefined
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What does Bile Esculin Agar contain?
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Beef extract, pancreatic digest of gelatin, esculin, bile, and ferric citrate.
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What is Esculin
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a glycoside composed of glucose and esculetin.
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Bile Esculin Agar inhibits growht of
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Gram positive organisms (except Grp D streptococci and enterococci)
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What is the indicator in Bile Esculin Agar?
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Ferric citrate
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Why does Grp D strep grow on Bile Esculin agar?
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it can hydrolyze esculin in presence of bile
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what color is formed when esculetin reacts with ferric citrate?
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Dark brown
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What is considered a positive in the Bile Esculin test?
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An organism that darkens the medium.
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What is the Bile Esculin test used to Identify?
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enterococci and members of the streptococcus bovis group all of which are positive.
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What is starch
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a polysaccharide made of of alpha-D-glucose subunits
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What are the 2 forms starch exists in?
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Linear (amylose) and branched (amylopectin)
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What do the enzymes alpha amylase and oligo-1,6-glucosidase do?
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Hydrolyse starch by breakin the glycosidic linkages
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What is a starch agar?
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Simple plated medium of beef extract, solublestarch and agar.
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What reagent is used to detect presence of starch around bacterial growth?
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Iodine
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Iodine reacts with starch and produces what color(s)
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Blue or dark brown.
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What was starch agar originally designed for?
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Cultivating Neisseria
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What is starch agar used for now?
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With pH indicators, it is used to isolate and identivy Gardnerella vaginalis
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What does species does starch agar differentiate?
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Cornynebacterium, clostridium, bacillus, bacteroides, fusobacterium and members of Enterococcus
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What does Eosin Methylene Blue (EMB) agar contain?
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Peptone, lactose, sucrose, eosin Y (dye) and methylene blue (dye)
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What do the sugars in the EMB agar do?
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provide fermentable substrates to grow fecal coliforms
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What do the dyes in EMB agar do?
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Inhibit growht of gram+ and produce dark purple complex with green sheen in acidic conditions
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How do slow growers and lack small amounts of acid appear on EMB?
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pink coloration
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What color are nonfermenters in EMB?
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Normal color or color of medium.
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What is EMB used to isolate?
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Fecal Coliforms
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Hektoen Enteric Agar (HE) is what type of medium and isolates what?
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Undefined medium. Isolate Salmonella nd Shigella from other enterics.
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What type of medium is MacConkey agar (MAC)?
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it is selective and differential.
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What does MAC contain?
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Lactose, bile salts, neutral red, and crystal violet.
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What do the bile salts in MAC do?
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Along with crystal violet, it inhibits growth of gram+ bacteria.
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Neutral red is a pH indicator. At less than __ it is ___. At more than__ it is _____.
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6.8, red
6.8, colorless |
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What color do lactose fermenters turn on MAC?
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a shade of red
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What is MAC used for?
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To isolate and differentiate membersof Enterobacteriaceae based on ability to ferment lactose.
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What does MAC without Crystal violet allow?
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Growth of gram+ cocci
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Describe Phenylethyl Alcohol agar
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Undefined, selective medium that allows growth of g+ organisms
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What is the active ingredient in Phenylethyl alcohol and how does it function?
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Phenylethyl alcohol, fn by interfering with DNA synthesis in Gram- organisms
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PEA is used to Isolate what?
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Staphylococci and streptococci from specimins.
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When Prepared with 5% sheep blood, PEA cultivates
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Gram positive anaerobes
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What is urea
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Product of decarboxylation of certain amino acids
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What can Urea be hydrolyzed to?
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Ammonia and carbon dioxide
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What enzyme hydrolyzes urea?
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Urease
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What members are considered rapid urease-positive organisms?
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Proteus, Morganella, and Providencia
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Why is Phenol red in urea broth?
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To expose increase in pH
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What color of Urea broth is positive within 24 hours? Negative?
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Positive is Pink
Negative is orange or yellow |
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What is Urea broth used for
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differentiate organisms based on their ability to hydrolyze urea with urease.
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What genus can be distinguished by using Urea hydrolysis?
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Proteus
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What is Casease
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An enzyme some bacteria produce to hydrolyze casein
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What is casein?
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Milk Protien that gives milk its white color
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Describe Milk agar
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undefined medium contaning pancreatic digest of casein, yeast extract, dextrose and powdered milk.
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What is the result of a positive innoculation?
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Secreted casease will diffuse into the medium around the colonies and create a clearing zone
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What is the casein hydrolysis test used for?
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Cultivation and differentiation of bacteria that produce casease
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What are the four phases of microorganism growth curve?
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Lag phase, Exponential (log) phase, Stationary phase, and death phase.
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Describe lag phase
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Cell synthesizing new components (replenish and adapt). varies in length
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Exponential (log) Phase
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Rate of growth constant. Population most uniform in phys and chem properties.
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Balanced growth
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During log phase. Cellular constituents made at constant rates relative to each other.
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Unbalanced growth
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rates of synthesis varies. conditions include change in nutrient level or env. condition.
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Stationary Phase
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Total number of VIABLE cells remains constant. May occur b/c reproductive rate is balanced by death rate.
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Reasons for entry into stationary phase
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Nutrient limited, oxygen limited, toxic waste accumulation, population density reached.
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Death phase (2 hypotheses)
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1. VBNC (viable but not culturable
2. Apoptosis |
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What does viable mean?
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Ability to reproduce
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Virulence
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abiliy to produce components that allow organism to become pathogenic
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pathogenicity
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Ability to produce symptoms of disease
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Generation time (doubling time)
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time required for population to double. varies from several min. to several days
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How do you count by spread plate?
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dilute plate with known amount on agar plate
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What are 2 methods of direct cell counts?
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Counting chambers, electronic counters, membrane filters.
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Types of viable cell counts
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plating methods (spread, pour)
membrane filtration |
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What does a Turbidostat do?
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Regulates flow rate of media through vessel to maintain a predetermined turbidity or cell density
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Extremeophiles
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grow under harsh conditions that would kill most others. make up of cytoplasic membrane.
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Fatty acid characteristics of Extremophile cytoplasmic membrane
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higher temp, f.a. more saturated.
Longer chains of FA |
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Water activity
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amount of water available to organisms
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What will reduce water activity?
|
Interaction with solute molecules (osmotic effect)
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pH formula
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-log [H]
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What pH range do acidophiles grow?
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0-5.5
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What pH range do neutrophiles grow?
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5.5-8
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What pH do alkalophiles grow?
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8-11.5
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Most acidophiles and alkalophiles maintain an internal pH near _________.
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Neutrality
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Most media contain buffers to do what?
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prevent growth inhibition.
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What are some ways thermophiles stabilize protein structure?
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-More H bonds
-more proline -chaperones |
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What stabilize DNA?
|
histone line proteins
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What are toxic products of oxygen reduction?
|
Superoxide radical
Hydrogen peroxide Hydroxyl radical |
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What protective enzymes do aerobes produce?
|
Superoxide dismutase (SOD)
Catalase |
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Barotolerant
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adversely affected bgy increased pressure, not as severe as nontolerant
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Barophilic orgnisms
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require or grow more rapidly in the presence of increased pressure (400 atm or more)
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What type of radiation is routinely used for killing?
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Gamma rays
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Why is UV light poor at killing?
|
It cannot penetrate plastic or glass.
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How is biofilm formed?
|
microbes reversibly attach to conditioned surface and release polysaccharides, proteins and DNA.polymr added
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Quorum sensing
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AHL or other signal molecule diffuse or att. to plasma membrane. enters cell. express target genes reg. fn
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AHL
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Acylhomoserine Lactone
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Sterilization
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destruction or removal of all viable organisms and viruses
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Disinfection
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Only to pathogens. Killing, inhibition or removal of pathogenic organisms.
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Sanitation
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Reduction of microbial population to levels deemed safe
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Antisepsis
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Prevention of infection of living tissue by microorganisms and viruses
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Antiseptics
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Chemical agents that killor inhibit growht of microorganisms when applied to tissue
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Antimicrobial agents
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agents that kill microorganisms or inhibit their growth and virus growth
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-cidal agents do what?
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Kill
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-static agents do what?
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inhibit growth
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good killing agent kills how much?
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10^5-10^6
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What contitions influence effectiveness of antimicrobials?
|
Population size
population composition Cenc. of a.m agent, duration, Temp, environment |
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What is physical method is highly effective for sterilizing?
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Heat
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Which physical method will not sterilize but will remove bacteria except mycoplasm and viruses
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Filtration
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What is the the best method of sterilization?
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Moist heat sterilization
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Thermal death time
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Shortest time needed to kill all microorganisms in a suspension at a T and defined conditions
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Decimal reduction time
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Time required to kill 90% of microorganisms or spores in a sample at a specific Temp.
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What is pasteurization used for?
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Milk, beer, and other beverages
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What was pasteurization originally used for?
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Grape juice
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What us pasteurization?
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kill pathogens and slow spoilage reducing total organisms present by controlled T below boiling
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What type of Sterilization oxidizes cell constituents and denatures proteins?
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Dry Heat. 160-170 for 2-3 hrs
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What does HEPA stand for?
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High-efficiency particulate air
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UV radiation is limited to what type of sterilization?
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Surface
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What penetrates deep into objects, destroys bac. endospores, not always viruses and used for ster/pas
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Ionizing ratiation
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What are the only types of aldehydes that will sterilize?
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Liquid aldehydes
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What is the hardest to kill?
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Spores
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What group is hardest to kill?
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Mycobacterium
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what will kill mycobacterium
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phenolics
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Where do phenolics come from?
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Coal tar
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Alcohols
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Bactericidal, fungicidal, not sporicidal. inactivate some viruses. Denature protein, dissolve membrane lipids
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What halogens are most used antimicrobials?
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Cl, F, I
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Which are most important halognes?
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F, Cl
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Iodine
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Skin antiseptic, oxidizes cell constituents, may kill spores
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Chlorine
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oxidizes cell constituents, used in water and houshold disinfectant. does not kill spores. can for carc. comp.
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What heavy metal has an affinity for gonnorhea?
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Silver nitrate 1%
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Detergents
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organic molecules with hydrophilic and phobic ends. act as wetting agents and emulsifiers
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Cationic detergents, benzalkonium Cl
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Kill most bacteria, not mycobacterium tuberculosis or endospores. safe but inactivated by soap/hard water
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Aldehydes
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sporicidal and chemical sterilants. combine with and incactivate nucleic acids and proteins.
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Chemotheraputic agents
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Chem. that can be used internally to kill ir inhibit growth of microbes
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Selective toxicity
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target microbe without harming host
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antibiotics
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Chemicals synthesized by microbes effective in controlling growth of bacteria
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What does most respiration involve use of?
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Electron transport chain
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Final electron acceptor in aerobic respiration
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oxygen
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final e- acceptor in anaerobic
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organic, or nitrates, sulfates, CO2, Fe3+
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What force is used to synthesize ATP?
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Proton motive force
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Fermentation
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uses endogenouis e- acceptor. Does not use ETC or generate PMF. ATP synth. by substrate level phsphrylton
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3 stages of Aerobic catabolism
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Polymers to momomers
oxidation and degrad. to pyruvate ox and degrad of pyruvate TCA |
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Amphibolic pathways
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Fn as catabolic and anabolic pathways
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3 common routes of breakdown of glucose to pyruvate
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Embden-Meyerhof Pathway
Pentose Phosphate Pathway Entner-Doudoroff Pathway |
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Embden-Meyerhof Pathway
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Occurs in cytoplasmic matrix
Most common path for glucose degradation to pyruvate in stage 2 of Aerobic resp. |
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Summary of Glycolosis
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Glucose+2ADP+2Pi+2NAD+
v 2pyruv+2ATP+2NADH+2H+ |
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Pentose Phosphate Pathway
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aka hexose monophosphate path
operate same time as glycolytic or Entner-Doud aerobic or anaerbic |
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Oxidation steps produce what
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NADPH for biosynthesis
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Pyrophosphate
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2 phosphate together can be used for energy in place of ATP Ex.Archea
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Entner Doudoroff pathway
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Not used in most prokaryotes
yield 1 ATP, 1NADPH, and 1 NADH per glucose molecule. |
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Where is prokaryotic ETC?
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plasma membrane
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Oxidative Phosphorylation
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ATP synthesized as result of electron transport driven by oxidation of chemical energy source.
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Theoretical and actual yield of ATP during Aerobic resp.
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Theoretical: 38
Actual: closer to 30 |
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Function of Krebs Cycle
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Reduction of high energy compounds
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Dissimilatory nitrate reduction
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use of nitrate as terminal electron acceptor
anaerobic red. of nitrate makes it unavail. for uptake |
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Denitrification
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Reduction of nitrate to nitrogen gas. In soil, causes loss of fertility
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How is ATP formed if oxidative phosphorylation does not occur?
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Substrate level phosphorylation
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Name some monosaccharides
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Fructose, Galactose
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Protease
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hydrolyzes protein to amino acid
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Deamination
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removal of amino group from amino acid. results conv. to pyruvate, acetyl CoA or TCA intermediate
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Stable, heritable changes in base sequence of DNA
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Mutations
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Permanant genetic change
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Mutant
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Induced mutations
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Changes caused by agents that directly damage DNA
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Types or causes of induced mutations (3)
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Base analogs
DNA-midifying agents Intercalating agents |
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Base analogs
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Chemotheraputic agents
structurally similar to normal mistakes occur when inc. into ggrowing polynucleotide chain |
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DNA modifying agents
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Alter a base causing it to mispair
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Intercalating agents
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Distort DNA to induce single nucleotide pair insertions and deletions
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What is the most used DNA modifying agent?
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Methyl-Nitrosoguanidine
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UV damage to DNA results in
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Formation of thyamine dimers
DNA can no longer serve as a template |
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Wild type
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Most prevalent form of a gene
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Forward mutation
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From wild type to mutant form
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Reverse mutation
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Mutant phenotype to wild phenotype
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Supressor mutation
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type of reverse mutation
occurs when secdond mutation is at a different site than the original mutation. |
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Point mutations (types)
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Silent, missense, nonsense, and frameshift.
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How are point mutations named?
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By if and how they change the encoded protein
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Silent mutation
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Change nucleoside sequence of codon, but not encoded amino acid
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Missense Mutation
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Single base substituion that changes codon fo one amino acid into codon for another
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Nonsense
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Converts a sense codon to stop codon
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Frameshift mutations
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results form insertion or deletion of one or two base pairs in the coding region of the gene.
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Conditional mutations
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expressed only under certain environmental conditions
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Auxotrophic Mutant
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Unable to make an essential molecule such as amino acid or nucleotide
Has cond. Phenotype |
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Prototroph
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wild type strain from which auxotrophic mutants arose
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Replica plating
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Screening technique for mutant deletion and selection. Used to detect auxotrophic mutants
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Some methods used for selecting mutants resistant to particular environmental stress
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UV light
Ionizing radiation Chemicals |
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Who created and used replica plating and when?
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Lederberg in 1952
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Carcinogenicity testing
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Based on observation that most carcinogens are mutants.
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What test is used to detect carcinogenic mutants?
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Ames test
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Discuss the Ames test procedure
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obtain culture. plate on plate with minimal medium and histidine. One plate with mutant one without. Innoculate at 37 revertants induced by mutagen number increase
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In Ames test, Describe in words the reversion rate comparison
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reversion rate in presence of suspected carcinogen is greater taht reversion rate in abscence of suspect carcinogen
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In Ames test, what is test mutagen often treated with?
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Mammalian liver extract
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Proofreading
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Correction of errors in base pairing made during replication
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How are proofreading errors corrected?
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By DNA polymerases
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Excision repair
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Corrects damage that causes distortions in double helix.
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Two types excision repair systems
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Nucleotide excision repair
Base excision repair |
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Photoreactivation is a method of what type of repair?
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Direct repair
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What is photoreactivation used for
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Directly repair thymine dimers
Thymines separated by photochemical reaction using visible light catalyzed by photolyase |
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What bacteria uses photoreactivation?
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E. coli
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How is direct repair of alkylated bases catalyzed?
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By alkyltransferase or methylguanine methyltransferase
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Mis match repair is what type of repair
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Excision repair
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What organism uses mismatch repair
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E.coli
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Discuss mismatch repair
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Mismatch correction enzyme scans newly made DNA for mismatched pairs. those pairs removed and replaced by DNA polymerase and DNA ligase
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What type of repair is SOS response?
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Inducible repair
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How many enzymes are involved in SOS response?
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21
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What is SOS response used for?
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To repair excessive damage tha halts replication, leaving many gaps. Highly error prone, used in life or death situation
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Discuss SOS response flow
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RecA protein initiates recombination repair
RecA also acts as protease destroys repressor protein thereby increasing productin of excision repair enzymes |
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Recombination is
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The process in which one or more nucleic acids are rearranged or combined tp produce a new nucleotide sequence
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How does Horizontal(lateral) Gene Transfer (HGT) occur in procaryotes?
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3 mechanisms. Conjugation, transformation, and transduction.
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What does HGT create?
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recombinants
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Transponons
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Segments of DNA that move about in genome. Can be integrated into different site. Also called jumping genes
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Insertion sequences
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Simplest transposable elements
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What type of DNA do plasmids have?
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Small circular DNA
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Episomes
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Small, autonomously replicating DNA molecules that reversibley integrate into host chromosome
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Conjugative plasmid Example
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F plasmid
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Transfer of genes between bacteria depends on
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Direct cell ot cell contact mediated by F pilus
Type IV secretion system Rolling circle replication of plasmid |
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F+xF- mating
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Copy of unintegrated F factor transferred to recipient and not integrate in host chromosome. Chromosomal genes not usually trans. from donor to recipient
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Donor HFr cell has what integrated into its chromosome?
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F Factor
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Complete copy of F factor usually not transferred due to
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Mating cells not attached long enough
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When does recipient cell become F+?
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When whole chromosome is transferred
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F' Conjugation
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Result when integrated F factor incorrectly leaves host.
Some left behind in host Some genes hve been removed along with some F factor. |
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F' x F-
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Yields 2 F'
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Competent cell
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on surface or associated has enzymes that can break down DNA into framents
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Uptake of DNA by a competent cell is followed by
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Incorporation of the DNA into the recipient cell's genome
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How can a cell be made more competent?
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By treating with Calcium
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Transduction
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Transfer of bacterial genes by viruses
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2 cycles a bacteriophage can carry out
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Lytic or lysogenic
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Lytic cycle
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Host cell destroyed causes cell to rupture
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Lysogenic cycle
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viral DNA integrate into host becoming a latent prophage
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Lysogen
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Host cell containing the prophage. Usually unharmed
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Lysogeny
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The relationship between prophage and host
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2 types of generalized transduction
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Generalized, Special
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When does Generalized transduction more frequently occur?
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During lytic cycle
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HFr mapping
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Used to map location of bacterial genes.
based that transfer occurs at constant rate |
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Interrupted mating experiment
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HFr x F- mating interrupted at various intervals. Order and timing of gene transfer determined
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