Kohlering/Serial Dilution/Streak/Enumeration
When streaking on an agar plate, when is it necessary to start over (discard current plate and grab new one)? Essentially, what must you make sure NOT to do?
When you pierce or puncture the agar.
If you focus your given bacterial sample on the slide after Gram staining and each cell is stained dark purple, is the organism Gram positive or Gram negative?
Gram-positive.
Your sample did NOT grow on MacConkey agar. Is your sample most likely Gram-positive or Gram-negative?
Gram-positive.
You got your motility broth back and you can only see growth down a very narrow tunnel in the center of your broth. Was your organism motile?
No.
What does a faint but intact and visible band on your gel indicate? Should you just ignore this?
It simply indicates that your sample had a low concentration. Do NOT just ignore it, as an intact band means that you got your desired result. The intensity/brightness/thickness should not scare you - it's okay if it's faint. As long as you can still see it, that counts as a result.
If each Eppendorf tube in a serial dilution has 1 mL of fluid TOTAL, how many microliters of the INITIAL, undiluted sample should be in the 10^-3 dilution, theoretically?
1 uL.
Is Staphylococcus aureus Gram-positive or Gram-negative?
Gram-positive.
What color should Staphylococcus aureus appear on Mannitol Salt Agar and why?
Yellow; Staph aureus is capable of fermenting mannitol. The agar itself selects for Staphylococcus because of it's high salt concentration.
What color should the cotton swab turn after the oxidase test IF your organism produced cytochrome c oxidase?
Violet to purple in seconds. If it's light pink or light purple, it's NEGATIVE.
You are given an image of a gel to analyze. You can see four wells: one ladder and three unknown samples. The bands on the ladder have their sizes clearly indicated. You are told that these are the results of a gel after loading the PCR product of a sample that had a target length of 450 base pairs into the unknown wells. What are you looking for on the gel to determine if the PCR worked for each of the samples in the unknown wells?
Look for an intact band between the bands on the ladder that are labeled 400 and 500 base pairs in length. Faint lines are okay and are still indicative of the PCR reaction working. If the lines are heavily smeared or nonexistent, then you can conclude that the PCR did not work for the sample in that well.
You are given the Petrifilm of a 10^-5 dilution sample. You count 62 colonies. How would you calculate how many CFU were in 1 mL of the original sample (assuming you pipetted 1 mL of diluted sample onto the Petrifilm and you started with 120 uL of the undiluted sample)?
62 x 10^5 x 8.333
If the majority of the cells in your sample are spherical-shaped and arranged in a single-file chain, what would the technical term for that arrangement be?
Streptococcus.
What is the difference in appearance on blood agar between organisms capable of beta hemolysis and those capable of alpha hemolysis?
Beta hemolysis: media around colony is fully transparent, lost its color.
Alpha hemolysis: Green or brown discoloration around colony.
Gamma hemolysis: NO hemolysis occurred (no color change at all, just see the colony growing as normal on the agar).
Which shape and arrangement is typical of E. coli?
Rod-shaped (bacillus) and single-celled (but can be found in pairs or short chains/clumps).
You got the results of your Sanger sequencing back after performing a PCR. What online resource would you use to find the species that your nucleotide sequence belongs to?
BLASTn (nucleotide BLAST).
Name the steps involved in properly Kohlering a microscope.
Focus using 4x or 10x lens, close field diaphragm completely, adjust condenser height to get the polygon with crisp edges (condenser all the way UP then slightly down - orange to blue on the edges), center circle, open field diaphragm to enlighten whole field of view.
NEVER touch WHICH lens with the immersion oil?
40x. ONLY use immersion oil on the 100x lens.
Is Staphylococcus aureus capable of producing deoxyribonuclease?
Yes.
How will you know if your organism tested positive on the catalase test?
Bubbles formed on your sample within seconds.
You are given a nucleotide sequence and you want to find the protein sequence that it likely corresponds to. You also want to be able to easily check the structure and function of the protein. Which resource would you use on BLAST and which database is best?
Blastx, UniProt/Swissprot database.
What function in Excel will calculate the standard deviation for your chosen data set?
=STDEV
Name the 5 crucial steps involved in Gram-staining. Go by the substance used in each.
Water + sample, dry, decolorizer to fix/glue cells. Crystal violet. Iodine/mordant. Decolorizer (drop by drop until it runs clear). Safranin (counterstain).
You are looking at the results of your phenol red broth test and you noticed that your sample turned bright yellow. There is also a clear/transparent bubble in the top of the inverted Durham tube in the vial. What do you know about your sample?
Your organism was CAPABLE of fermenting the carbohydrate in the medium AND it is heterofermentative (gas was produced along with acid).
If your organism is capable of decomposing tryptophan to indole, what color should the layer of fluid on the top of your motility medium be after you add Kovac's reagent?
Dark red.
Which formula is used to manually calculate the primer's melting temperature (when the primer melts off the DNA)?
(G and C x 4) + (A and T x 2)