BIOL107 Study Guide - Final Guide: Eyepiece, Betalain, Fluorescence Microscope

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silverlouse330

29 Nov 2016

School

University of Alberta

Department

Biology (Biological Sciences)

Course

BIOL107

Professor

Kirstin Veugelers

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BIOL107 Full Course Notes

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Document Summary

Lab 1: introduction, microscopes, scientific method, and osmosis. Cell biology is the study of how cells or parts of a cell function. A pure culture is an isolated group of genetically identical cells. They can easily be contaminated with another cell type as microorganisms are everywhere in the environment. The chances of contamination can be reduced by using aseptic (sterile) technique. One aspect of aseptic technique is to work in an area with a lowered risk of contamination. A colony is a small mass of genetically identical cells. In lab 1, we took imprints of cellphones in a solid medium. A medium is a solution or solid that contains nutrients cells can use to grow and divide. Individual cells transferred to the plate will grow and divide, eventually forming colonies. Gadgets were swabbed with a sterile medium, then the swab was incubated in liquid medium to allow the microorganisms to grow and multiply.

Related Questions

Considering the cultures to be used for inoculating each medium in this exercise, how many microbial types should you expect to see in/on each medium?

Select one:

a. 3

b. 0

c. 1

d. Several

e. 2

Which of the following is common to minimize airborne contamination?

Select one:

a. Make someone else do the experiment

b. Perform the experiment in a windy room

c. Keep a microscope nearby

d. Breathing in sample

e. Hold open tubes at an angle

How can a hot sterile flame loop be cooled quickly to use in the next streak?

Select one:

a. Put in sterile portion of agar plate

b. Blow on it

c. Stick the loop in ice

d. Set the loop on the lab bench

e. Pour tap water on the loop

True or false: Most colonies can been seen without aid (naked eye), and most single cells cannot be seen without a microscope.

Select one:

True

False

An isolated, single colony forms after incubation of a:

Select one:

a. Virus (viral particle arrangement in some cases)

b. Single cell (cell arrangement in some cases)

c. Sterile agar plate

d. Sterile flame loop

e. Liquid mixed culture

These are commonly used for growing stock cultures that can be refrigerated for several weeks:

Select one:

a. Broth tubes

b. Cotton swabs

c. Gym socks

d. Agar slants

e. Petri dishes

Beginning at the base of the slant surface and gently moving the loop back and forth as you withdraw is called:

Select one:

a. Slant mixing

b. Fishtail inoculation

c. Zig-zag streaking

d. Squirrel inoculation

e. Quadrant streaking

If you are going to mix a broth culture prior to transfer, what is a safe way to do this without special equipment?

Select one:

a. Tap the tube with your fingers

b. Vortex it

c. Use a inoculating loop

d. Stick the tube in the outer cone

What is the primary purpose of the streak plate method?

Select one:

a. Quantify CFUs

b. Isolate an individual colony

c. Quantify the cell count of a mixed culture

d. Inoculate a culture on a plate

e. Confirm the identity of an unknown genus

Please answer questions at the end.

In this lab, you will add different volumes of âcell-free supernatantâ (CFS) to fresh cultures of V. harveyi and observe bioluminescence. The CFS is prepared by culturing V. harveyi in liquid medium overnight, then separating the cells from the supernatant by centrifugation.

MATERIALS â¢ overnight culture of V. harveyi in AB medium â¢ fresh AB medium â¢ 1 microcentrifuge tube â¢ 3 glass culture tubes

PROCEDURE

1. This experiment will be performed with your lab group. Label the microcentrifuge tube and 3 glass cultures tubes with your group number. Also, label the 3 glass culture tubes as follows: âMedia only,â âCells+fresh media,â and âCells+CFS.â

2. Using aseptic technique, pipet different volumes of fresh AB medium into the glass tubes as follows: 1 mL in the âMedia onlyâ tube, 800 ul in the âCells+fresh mediaâ tube, and 500 ul in the âCells+CFSâ tube.

3. Pipet 500 ul of the overnight culture into a microcentrifuge tube and centrifuge at maximum speed (>13,000 x g) for 5 min to pellet the cells.

4. After centrifugation is complete, transfer 300 ul of the cell free supernatant (CFS) from the microfuge tube to the glass tube labeled âCells+CFSâ. Be careful to avoid the cell pellet.

5. Remove the remaining supernatant from the microfuge tube using a micropipette and discard as culture waste.

6. Add 500 ul of fresh AB medium to the cell pellet in the microfuge tube and pipet up and down to resuspend until no clumps are visible.

7. Add 200 ul of the cell suspension to the âCells+fresh mediaâ tube and 200 ul of the cell suspension to the âCells+CFSâ tube. Note the time.

8. Place the 2 glass tubes containing cells in the 30 degree incubator/shaker.

9. After 1 hour and 2 hours, observe your tubes next to the âMedia onlyâ tube in a dark room. You will need to let your eyes adjust for at least 30 seconds (count to 30 slowly in the dark) before you can evaluate whether the cultures are luminescent or not. Also, make sure you try swirling the tubes in the darkroom and observe the tubes for a minute after you have stopped swirling. After the observation at the 1 hour time point, put your tubes back in the incubator shaker until the 2 hour time point.

10. Note your observations. At the end of the lab, place your tubes in the tube waste.

Questions

1. What were the expected results? Explain the rationale of the experiment. If you didnât obtain the expected results, hypothesize why not and describe how the experiment could be improved.

2. What are differences between luminescence and fluorescence?

3. Why do V. harveyi cultures glow more brightly when swirled vigorously? Be specific.

4. Imagine you have a mutant V. harveyi strain that canât synthesize autoinducer and another mutant V. harveyi strain that makes a defective luciferase. If you streaked both strains near (but not touching) each other on the same plate, would you expect to see light produced? If so, by which strain, where, and why?

5. You have two identical tubes of wild-type V. harveyi (with the same cell concentration in each tube) in fresh media. To one tube, you add CFS from a light-producing V. fischeri culture, and to the other tube you add an equal volume of fresh media. If both tubes of V. harveyi have the same light production, what can you conclude? Explain.

6. Many biomedical research labs study quorum sensing in V. fischeri or V. harveyi. How is studying light production in marine bacteria relevant to human health? List at least 3 different examples.

7. Why do you think it is important for bacteria to be able to control the expression of some genes based on the number of individuals present?

graylion297

In a quadrant streak, why do you intersect the previous lines and not the ends of the streak?

Select one:

a. More cell amount variability at ends, usually higher cell population

b. Keep streaks in the middle of the plate

c. Prevent cell death

d. To maintain a mixed culture

e. To count CFUs

How should you hold the inoculating loop?

Select one:

a. Maintain it at an angle at all times

b. In a test tube rack at all times

c. Like a pencil

d. Balanced on top of the tube

e. With dish gloves

Which of the following is used in the spread plate and not the streak plate method?

Select one:

a. Glass spreading rod

b. Flame loop

c. Agar plate

d. Bunsen burner/sterile utensil

e. Starting culture to sample

What would likely happen if the inoculum was not spread adequately over the agar surface?

Select one:

a. Contamination

b. Degradation

c. Sterilization

d. Uneven distribution

e. Cells will not grow

You have a mixed culture of unknown bacteria and want to identify one of the species by several subsequent experiments (needing multiple colonies of a pure culture). You do a quadrant streak and transfer to a sterile liquid medium. What next step makes the most sense?

Select one:

a. Perform a Gram stain

b. Note all colony morphology characteristics

c. Use the spread plate method or new streak plate on an isolated colony of the plate

d. Quantify the cells

e. Repeat using the other two methods of isolation

What would likely occur if the flame loop was not sterilized between streaks on the quadrant streak method?

Select one:

a. Bacteria will not stick to the loop

b. Virus will be introduced

c. Flame loop will be too hot

d. The sample will be contaminated

e. Subsequent streaks would have more colonies

Why is the glass rod in the spread plate method put in alcohol and ignited instead of sterilized like a flame loop?

Select one:

a. Glass oxidizes

b. Glass melts

c. Glass takes longer to cool

d. Glass burns

e. Glass is more sterile

BIOL107 Study Guide - Final Guide: Eyepiece, Betalain, Fluorescence Microscope

BIOL107 Full Course Notes

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