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81 Cards in this Set
- Front
- Back
Bacterial transformation?
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process by which competent bacterial cells pick up DNA from the environment & make use of the genes it carries?
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How is E. Coli made "competent"?
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made artificially via CaCl2 transforming solution and heat shock
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What are plasmids?
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when a bacteria naturally contain one or more small circular pieces of DNA
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What causes bacterial resistance?
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the transmission of plasmids
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what is an operon?
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structural and functional genetic units of prokaryotes
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What is the basic structure of an operon?
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operon minimally induces a promoter site (for binding RNA polymerase) and two or more structural genes coding for enzymes in the same metabolic pathway
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What operon did we use in class?
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the arabinose operon
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What are the four components of the arabinose operon?
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the promoter PBAD and three structural genes (araB, araA, and araD) that code for enzymes used in arabinose digestion
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What part of the arabinose operon functions during lack of arabinose and how does it act?
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araC (a DNA binding protein) prevents RNA polymerase from binding to the promoter, so transcription cannot occur – the switch is “off.”
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How does araC act in arabinose presence?
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- When arabinose is present, it binds to araC and changes its shape so RNA polymerase can bind to the promoter and transcribe the genes – the switch is “on.”
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What is pGLO's use in our experiment?
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replaces the structural genes (araB, araA, and araD) with the GFP gene
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What antibiotic gene does pGLO contain?
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bla
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What is bla and what does it produce? What does that enzyme do/
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It is an gene coding for ß-lactamase, which hydrolyzes certain antibiotics, including penicillin and ampicillin
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A dish contains E. coli crossed with P. aeruginosa. It is a ...
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mixed culture.
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A dish contains only one species...
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pure culture
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What does the streak plate tenchinuque accomplish?
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to identify species from a mixed culture (see Fig 1-24 a-d)
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The zigzag technique is used when a sample does not have a high cell denisty and is with pure cultures. Why is this techinque used?
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The zigzag technique is one that is meant to harbor individual colonies. These colonies can then be isolated for growth in a new medium.
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What are the major components of Fluid Thioglycollate Medium.
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Sodium theioglycollate, L-cystine, pancreatic digest of casein, resazurin
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What is the main purpose of Fluid Thioglycollate Medium?
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To provide an anarobic environment with nutrients for growth.
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In Fluid Thioglycollate Medium, which components provide "food"?
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the yeast extract and casein (from pancreatic digest)
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In the fluid Fluid Thioglycollate Medium, which keeps the medium anaerobic?
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the thioglycollate and L-cystine
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What is resazurin? There are two colors it takes. Name the colors and which state is associated with it.
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Resazurin is an indicator.
pink when oxizidized colorless when reduced |
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What two layers exist in the Fluid Thioglycollate Medium.
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The top layer is aerobic. The bottom layer is anaerobic. It is a gradient between them.
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How does the top layer of Fluid Thioglycollate Medium form?
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Initially, the whole medium lack O2 from autoclaving. Afterwards at room temperature, O2 diffuses into the top layer without fully penetrating the bottom layer.
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What is autoclaving?
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Subjecting items to conditions of high pressure and high temperature sterilization.
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This needs oxygen to grow.
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aerobe
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An organism that grows in the absence of free oxygen
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anaerobe
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I (bacteria) grow better with oxygen.
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faculative anaerobe
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Some O2 is OK is me (2 - 10%), but too much kills
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Microaerophile
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What are the unites of microbial cell denisty?
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cells/mL
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How do we indirectly measure microbial cell density?
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By use of a standard plate count.
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What is serial about serial dillutions?
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It invovles multiple dilutions, each dillution from the PREVIOUS dillution, thus compounding the result to even as small as a millionth of original density.
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Process of counting number of colonies a portion of sample produces when spread onto an agar plate.
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standard plate count
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Why is it important to reduce cell density using serial dillutions?
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1. yields countable plates
2. math can be used to find OCD (original cell denisty) |
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What is the formula for DF?
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Dilution factor: volume of broth aded to diluent/
(total volume after the broth was added) |
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What is OCD formula?
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colunies counted/
[(Volume plated)(DF)] |
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Phenol red is a ...
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indicator
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Phenol red is red in pH of ...
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8 or higher
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Why would phenol red turn yellow?
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fermentation of carbohydrate (glucose, lactose, sucrose) creates acidic conditions. At pH below 6.6 the solution turns yellow.
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What does deamination do?
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Breaks down amino acids (duh). Main point: creates NH3, raising pH making solution red in phenol red presence.
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In PR test, how do we know if gas production occured?
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A bubble is found in the tube.
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If fermentation occurs in the PR test, there is a gas bubble in the ...
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...Durham tube.
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Which tests analyses pH change from mixed acid fermentation?
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Methyl Red Test
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What colors occurs in the Methyl red test and at what pH?
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pH lower than 4.4 - red
pH higher than 6.2 - yellow pH between - orange |
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An organism in the methyl red test produces no acid. The color of the test is ___ and the organism does not use _____
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1. yellow (orange maybe, inconclusive
2. mixed acid fermentation |
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What is the main purpose of Voges-Proskauer Test?
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It tests for glucose use with the end product of acetoin
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What is acetoin? (IUPAC name)
Is it neutral, acidic or basic? |
(2,3 - butanediol fermentation )
-neutral |
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A student got a clear solution for the Voges-Proskauer test. What does he conclude?
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The bacteria may use glucose but without the endproduct of acetoin.
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Name 2 oxides toxic to organisms?
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hydrogen peroxide, superoxide radical
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What protein produces toxic oxides in bacteria?
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flavoprotein
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The flavoprotein is a part of the ...?
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electron transport chain
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What does catalase do?
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It breaks down hydrogen peroxide to water and O2
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What breaks down superoxide radical?
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superoxide dismutase
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What is superoxide radical converted?
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hydrogen peroxide
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A positive catalase result means one sees...
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gas due to hydrogen peroxide being added
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Cytochrome c oxidase transfers what to what
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It transfers electrons to oxygen in some ETC
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Tetramethyl-p-phenylenediamine transfers what to what
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electrons to cytrochrome c oxidase
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What is the purple from the Oxidase test due to?
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It is from the tetramethyl-p-phenylenediamine transfering electrons to cytocrome c oxidase, thus confirming its presense.
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If citrate is the only source of carbon for the bacteria, the medium turns ____ instead of _____
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blue instead of green
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What is the dye in the citrate test? What pH makes it blue.
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Bromthymol blue is blue at a pH of 7.6
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What is citrate converted to create a blue 7.6 pH environment?
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ammonia and ammomium hydroxide
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Nitrate reductase is found in many gram ______ bacteria. It converts what to what? (nitrate reduction question)
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negative .... Nitrate (NO3-) to nitrite (NO2-) (NH3, NO, N2O also acceptable, as long as nitrate is inital reactant)
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What is denitrification?
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When Nitrate is converted to N2 (molecular nitrogen)
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Why should the fermentation (PR tests) test done before the Nitrate reduction test?
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Both fermentation and denitrification create air in the Durham tube. To conclude denitrification, one must eliminate fermentation as the product.
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There is no gas bubble after initial nitrate reduction test. You conclude...
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denitrifcation is not present. Nitrate reduction can still exist.
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There was no bubble for Nitrate reduction test. You add ____ and ____ to test for ______. A red solution meant your test was _____.
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nitrate reagent a;
nitrate reagent B; nitrite; positive |
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There was no bubble for initial nitrate test. You add two reagents. The solution remains clear. You then add ____. You notice a clear solution. You conclude what?
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Zinc. You conclude that nitrate was reduced to either NH3, NO, Or N2O or other nongaseous nitrogenous compound.
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There was no bubble for initial nitrate test. You add two reagents. The solution remains clear. You then add ____. You notice a red solution. You conclude what?
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zinc; Nitrate is not reduced by organism
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Decarboyxlation broth contains what indicator?
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Bromcresol purple
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What part of the decarboxylation broth promotes fermentation?
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The mineral oil seals the broth from external oxygen, thus creating conditions for fermentation.
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The decarboxylation test is performed, the student gets a purple solution. This happened because the organism produced _____ the enzyme causing a build up of ______ ___ ______.
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decarboxylase
alkanline end products |
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Decarboxylation occurs toa specific ____ _____.
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amino acids (different organisms will decarboxyalize different a. a's)
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The blue result from decarboxylation occurs when the pH is above ___.
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pH 6.8
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A student notices a yellow solution in the Decarboxylation test. He can conclude two things. What are they?
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1. The test is still in progress and will later be positive when the organism makes decarboxylase
2. The organism does not produce the approiamte decarboxylase. |
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Decarboxylases break down what functional group of a specific amino acid?
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the COOH
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The phenylalanine deaminase tests for an organisms to catalyze the ____ functional group of a specific ____ ____.
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NH3;
amino acid |
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A student notices a ____ color change for a positive phenylalanine deaminase test.
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green
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What familiar indicator is used in the urease test?
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phenol red
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The substance urease will break down _____. Urease creates a basic environment by producing NH3/NH4/OH
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urea
NH3 |
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A student performing the urease tests notices a _____ slant. He assumes the pH is above 8.4 and the enzyme _____ is present.
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pink
urease |
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If the urease test is negative, it will be pink/yellow/red/orange.
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Yellow. Pink or organge is considered positive, weak positive respectively
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