micro midterm

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Francis Marion University *

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2420

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Biology

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Jul 2, 2024

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MICR 3051 General Microbiology Lab (Spring 2023) Midterm Laboratory Exam Study Guide Labs1-5 Objectives: 1. Be able to distinguish between safe and unsafe microbiology laboratory practices. Safe: o Wear lab coat o Use goggles when working with/around heat (Imao) o Use aseptic technique o Hairup Unsafe: o Not wearing a lab coat Honestly, I feel like any questions about this will be rather easy to figure out what is safe and what isn’t safe lmao 2. Know the parts of the microscope and their functions. = = 0 Eyepiece lens (ocular lens) [fj what you look through [fj 10x Tube 2] connects the eyepiece to the objective lens Arm [?] connects the tube to base Base (2] bottom of the microscope Iluminator (7] light source Stage w/stage clips [?] platform where slides are placed Revolving nosepiece (2| holds the objective lenses and is rotated casily Objective lenses 2] 4x, 10x, 40x, 100x, what is used to see the specimens at varying magnifications Condenser lends (2] focuses light on the specimen Diaphragm or iris [?|] rotating disc under the stage with different-sized holes to vary the intensity and size of the cone of light Know how to use the microscope (adjusting for proper viewing, getting an image into focus, using the oil immersion lens) and how to take care of a microscope. Only use the coarse adjustment in the 4x and 10x magnification lens Use the fine adjustment on the 100x magnification lens Clean off the lenses with lens paper before and after use. Be sure to use a different piece of lens paper when cleaning the 4x and 10x if 100x was cleaned first — aka avoid spreading oil from 100x to the 4x or 10x. Always focus the microscope with the 4x first, then move to the 10x and focus again When going to 100x, you need oil immersion o Flip the rotating nosepiece to between the 4x and 100x

o o Add one drop of immersion oil to the slide o Move the 100x on top of the slide o Do NOT let the immersion oil touch any of the other slides When placing microscope into the cubbies, be sure to have the eyepiece facing outward towards you o Carry it using on hand on the base and one hand on the handle Describe the aseptic technique and understand why it is important. The aseptic technique uses practices and procedures to prevent contamination from pathogens — different techniques are applied to do this. Designed to provide a barrier between the microorganisms in the environment from the pure culture. It’s important because, without it, any of our tests (stains, biochemical tests, etc) could be contaminated and give us false results. It also protects us (the people) from being contaminated with bacteria we work with in the lab. Define and describe culture media. Culture media is a gel or liquid (or semisolid) that contains nutrients and is used to grow bacteria or microorganisms There are multiple different types of culture mediums that are different for what may be grown on them o Sclective o Differential o Enriched Compare and contrast how to make a bacterial smear from liquid media and solid media. You do not need to add anything to the slide when starting with liquid media. Using an aseptically clean inoculation loop, get a loop full of the liquid media and culture and spread it thinly on the plate. Be sure to shake the liquid media to ensure the bacteria is thoroughly mixed throughout. Solid media, however, will require a loopful of water to be added to the slide before adding the culture onto it. Be sure to spread this one evenly as well or else it will be hard to visualize on the microscope. Explain the importance of heat fixing and air-drying specimens on slides. Heat fixing the bacteria on the slide does: o Kills the bacteria o Firmly fixates it on the slide (less of a chance of it moving while staining or blot drying o Allows for the bacteria to take the stain better Aka it stains better when heat-fixed Air-drying specimens is important for: o Allows for all the water to be removed, when this occurs that means there 1s no chance for water to boil and destroy the bacterial cells during the drying process Compare and contrast simple staining and differential staining. Simple staining is done with the use of one singular stain

= & EE S o =] 11, Differential staining is done with the use of 2 different stains (and a decolorizing step) to show the difference between 2 different bacteria Recognize the different bacterial morphologies. Cocci — spheres o Tetrad — four in a cube shape o Sarcinac — cight in a cube shape o Diplococci —2 o Streptococci — in a chain o Staphylococci — clusters of cocci Bacilli — rods o Coccobacilli — short rods o Diplobacilli — 2 o Palisades — hooked together at 1 end Spirochetes Vibrio — comma shaped Filamentous Spiral . Compare and contrast gram-positive and gram-negative cells (what colors they stain using the Gram stain and how cell wall structure determines how they stain). Gram-positive: o A thick layer of peptidoglycan in their cell wall o Stain purple because of it The peptidoglycan shrinks in the decolorizing step, trapping the CV-I1 complexes in it Gram-negative: o A thin layer of peptidoglycan and an outer membrane o Stain pink because of that During the decolorizing step, outer membrane breaks down and the peptidoglycan layer is too thin to keep hold of the CV-I complexes, leaving it colorless so the counterstain of safranin can stain it pink Know all of the steps and stains used in a Gram stain. Know the purpose of cach stain and how bacteria look at cach step. Know which stain/reagent is the primary stain, counterstain, mordant, and decolorizing agent. Crystal Violet o Primary stain o Stains both cells purple Gram’s lodine o Mordant o Forms CV-I complexes o Both cells still purple Alcohol

o Deccolorizer o Gram-positive — purple o Gram-negative — colorless Safranin o Counterstain o Gram-positive — purple o Gram-negative - pink . Understand the purpose of a streak plate and how you would do one. The streak plating technique is done to get a single, isolated colony of a bacterial culture you’re studying Steps: o On one quadrant of the agar plate [?] inoculate it o Flame the loop and turn the plate o Drag loop through quadrant one and make one line, then make 2 more lines not touching quadrant one anymore underneath it Flame the loop Drag loop through all 3 lines in quad 2 and do the same thing as said above Flame loop Drag loop through all 3 lines from quad 3, and do the same thing as before Also, make random shapes in the center to get rid of the excess left on the loop (do not touch anything) O 0O O O . Define pure culture and CFU. Pure culture 7] lab culture containing a single species of an organism 2] all come from a singular organism that split to create daughter cells CFU ) colony forming unit (number of visible colonies on a plate) . Describe the growth of bacteria in various types of media (cultural characteristics) using the proper microbiological terms. Plate: o Size: Small Medium Large o Shape: Circular — any round colony regardless of type margin Irregular — not circular, may be spreading Punctiform — forming pinpoint colonies (multiple small colonies around cach other) Rhizoid (root-like) — elongated and branching, looks like tree roots o Color/pigmentation: Some bacteria produce color wen grown in the medium, depends on the bacteria and the media — probably the easiest one to

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