Also write M. As demonstrated, use a sterilized inoculating loop to pick up one M. Spread the bacteria over approximately a quarter of the plate, edge to edge. Consider this step 1. Flame the loop and cool it in the agar.
Overlap the step 1 streak times to pull out a reduced number of bacteria, and spread them out down the side of the plate. Consider this step 2. Overlap the step 2 streak times and spread over the surface. Continue this process, flaming the loop in between each step, until the entire surface of the agar plate is covered. After performing this with the M. Place the streak plate subcultures in an incubator at the temperature and time specified by your instructor.
Obtain one slant tube containing TSA, and label it using a small piece of tape with your name and culture name M. Using a sterilized inoculating loop , pick up a bacterial colony or piece of a colony from the surface of the plate culture of M. Place the slant subculture in an incubator at the temperature and time specified by your instructor. Obtain one stab tube containing semisolid TSA, and label it using a small piece of tape with your name and culture name E.
Using a sterilized inoculating needle , pick up a bacterial colony or piece of a colony from the surface of the plate culture of E.
Withdraw the needle carefully and try to remove it by following the same stab line that you made pushing the needle down. Place the stab subculture in an incubator at the temperature and time specified by your instructor. As you will learn, bacteria have preferred growth temperatures where their reproduction rate is the greatest.
This lab is equipped with incubators set at either temperature. How long you plan to leave your cultures in an incubator should also be a consideration.
Growing cultures at the higher temperature may speed their rate of growth, but it also causes dehydration of the media and an earlier demise to the bacteria in the culture. Bacteria that grow best at room temperature should always be incubated at room temperature, and growth may take a little longer.
It is important that you read the project description in the next chapter so that you understand the goals and the scope of the project. To begin, you will take a sample from your skin. Your first decision will be what part of your skin do you want to sample? Note: ONLY external skin surfaces are permitted.
Obtain a sterile swab and a tube of sterile distilled water, and label a TSA plate with your name and the date. Remove the wrapping from the swab and soak it in sterile water, using aseptic technique. Rub the wet swab back and forth firmly over the area of skin you have chosen to sample. Then rub the swab over approximately a third of the surface of the TSA agar plate. Sterilize an inoculating loop, and complete the rest of the streak plate pattern using the loop.
Incubate this plate at room temperature for up to a week. After incubation, look to see if isolated colonies have developed on the plate. If there are no colonies or no isolated colonies, you will need to make another streak plate with the advice of your instructor on how to proceed. If there are isolated colonies, transfer the plate to the refrigerator. From this plate you will ultimately choose one single colony and prepare a pure culture. To complete the lab , the bacteria in the cultures have to grow.
Cloudiness of broth before incubation Predicted appearance of broth after incubation Actual appearance of broth after incubation. Look at the streak plate subcultures that you made. Conduct a self-assessment of how well you performed the technique. What you hope to see are individual colonies, well separated from each other. On the streak plate of the mixed culture, you should be able to see two distinctly different types of colonies.
Streak plate of the mixed culture :. Look closely along the stab line in the media in the tube. For the purpose of gelling the microbial culture, the medium of agarose gel agar is used. Agar is a gelatinous substance derived from seaweed. A cheap substitute for agar is guar gum, which can be used for the isolation and maintenance of thermophiles. Microbiological cultures can be grown in petri dishes of differing sizes that have a thin layer of agar-based growth medium.
What factor allows organisms such as planarian worms and plants to regenerate new individuals from fragments of parent organisms. What kind of bacterium cannot tolerate oxygen. In what cycle of viral replication does the virus destroy the host cell.
How is yeast used. What are the five main groups of arthropods. What do Amoebas use for movement. How are algae different from plants. How do cells control osmosis. What is cytoplasm. Where is ATP often produced. What causes water to be a polar molecule. Q: How do the pure broth culture differ from the slant culture? Write your answer Related questions. Can a Pure culture prepared from mixed broth or mixed agar slant? Can a pure culture be prepared from a mixed culture broth? Can pure culture be prepared directly from a mixed broth culture?
What is a pure broth culture? Can you determine whether a broth culture is pure by visually inspecting it without a microscope? What type of culture technique would be used to obtain a pure culture? What is a pure culture? Can you determine whether a broth culture is pureall one species by visually inspecting it without a microscope?
Why is it important that the Tryptic Soy Agar slant be inoculated with a pure bacterial culture instead of a mixed culture? Probably the most visual characteristic is pigmentation color. Some microorganisms produce pigment during growth and are said to be chromogenic. Often, however, formation of pigment depends on environmental factors such as temperature, nutrients, pH and moisture.
Pigments can be divided into two basic types: water-insoluble and water-soluble. If the pigment is water-insoluble see Fig. As a result, the colonies are pigmented but the agar remains the normal color. If the pigment is water-soluble as in the case of Pseudomonas aeruginosa see Fig. Both the colonies and the agar will appear pigmented. Below is a list of several common chromogenic bacteria:.
See Fig. Remember to label all tubes with a wax marker. When streaking the agar plates, use either of the patterns shown in Fig. This procedure is termed streaking for isolation and has a diluting effect. The friction of the loop against the agar causes organisms to fall off the loop.
Near the end of the streaking pattern, individual organisms become separated far enough apart on the agar surface to give rise to isolated single colonies after incubation. Incubate all the tubes and plates inoculated with B. Place the tubes in your dedidated test tube rack. Incubating the plates upside down prevents condensing water from falling down on the growing colonies and causing them to run together.
Store your test tube rack on your incubator shelf when not in use. In order to illustrate that microorganisms are all around us and to demonstrate the necessity for proper aseptic technique, contaminate three Trypticase Soy Agar plates as follows:.
Remove the lid from the first agar plate and place the exposed agar portion in or out of the building for the duration of today's lab. Replace the lid, label the plate "air" , and incubate it upside-down at room temperature. Do this plate first. Using a wax marker, divide a second petri plate in half. You and your partner both moisten a sterile cotton swab in sterile water.
Rub your swab over some surface in the building or on yourself. Use this swab to inoculate your half of the second agar plate. Label the plate and incubate upside-down at room temperature. With a wax marker, divide a third petri plate into quartersand label as shown in Fig. On your half of the plate, first rub the fingers of one of your gloved hands over your "glove" quadrant.
Remove that glove and rub your fingers over your "fingers" quadrant. Your partner will do the same on his or her half of the plate. Do this plate last. Draw and describe the growth seen in each of the four broth cultures. Observe the growth in the slant cultures and stab cultures for pigmentation and purity. Using the terms in the Appendix A, compare a single colony of B. Use a hand lens or a dissecting microscope to magnify the colony.
Observe the results of the three "contamination" plates and note the differences in colony appearances. Observe the demonstration plates of chromogenic bacteria and state the color and water-solubility of each pigment. After completing this lab, the student will be able to perform the following objectives:.
Using an inoculating loop, demonstrate how to aseptically remove some inoculum from either a broth tube, slant tube, stab tube, or petri plate, and inoculate a sterile broth tube, slant tube, stab tube, or petri plate without introducing outside contamination. Label all tubes and plates and place them on the shelf in the incubator corresponding to your lab section.
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