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81 Cards in this Set

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