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How do you grow bacteria colonies?

 

*Recommended for Grades 8 to 12

Introduction: (Initial Observation)
Bacteria are very small micro organisms that can not be seen with a naked eye.  So we can not see them swimming in water. The other problem is that often millions of bacteria are condensed in a small spot. 

To see bacteria, you grow them in a nutrient agar plate so each bacteria will become a colony of thousands of bacteria. Then we can see them as a bacteria colony.

In this project you will perform experiments grow bacteria colonies.

Information Gathering:
Gather information about your project. If you are a basic or advanced member of ScienceProject.com, your project advisor may prepare the initial information that you need and enter them in this section. In any case it is necessary for you to read additional books, magazines or ask professionals who might know in order to learn more about the subject of your research. Keep track of where you got your information from.

Bacteria are single-celled micro organisms that play an important role in many ecosystems. Bacteria vary in terms of morphology, oxygen and nutritional requirements and motility. Bacteria may be free-living, live on dead or decaying organic matter (saprophytic), or pathogenic in plants or animals. Bacteria reproduce by fission. 

Question/ Purpose:
What do you want to find out? Write a statement that describes what you want to do. Use your observations and questions to write the statement.
Identify Variables:
When you think you know what variables may be involved, think about ways to change one at a time. If you change more than one at a time, you will not know what variable is causing your observation. Sometimes variables are linked and work together to cause something. At first, try to choose variables that you think act independently of each other.
Hypothesis:
Based on your gathered information, make an educated guess about the answer to your question or the result of your experiment. 
Experiment Design:
Design an experiment to test each hypothesis. Make a step-by-step list of what you will do to answer each question. This list is called an experimental procedure. For an experiment to give answers you can trust, it must have a "control." A control is an additional experimental trial or run. It is a separate experiment, done exactly like the others. The only difference is that no experimental variables are changed. A control is a neutral "reference point" for comparison that allows you to see what changing a variable does by comparing it to not changing anything. Dependable controls are sometimes very hard to develop. They can be the hardest part of a project. Without a control you cannot be sure that changing the variable causes your observations. A series of experiments that includes a control is called a "controlled experiment."

In order to grow bacteria, you will need culture media, plates or petri-dishes and some laboratory supplies and incubator. 

Culture Media: Culture media is a moist or liquid matter that contains nutrients for bacteria. Almost any nutrient food may be considered a culture media for general bacteria, however if you want to grow a specific bacteria or prevent growing some other bacteria, you will need to use a fine tuned recipe for your culture media. 

Chicken broth and beef broth are among nutrients that most bacteria like. In some recipes you may also add some mushroom extract. Sugar can also be added to most culture media. Small amounts of some minerals such as potassium phosphate and calcium carbonate may also be added to the culture media. Note that there are many foods that are good for growing bacteria, but they are not good as culture media. For example bacteria can easily grow on milk, but milk is not a good culture media because it will change by the activity of bacteria. Part of milk will solidifies when bacteria produce acids. A good culture media must be clear and must remain liquid and should not easily change pH. If we need to solidify our culture media, we use agar to do that. Agar is a gelatinous substance that is extracted from sea weeds. If we need to grow bacteria for the purpose of identification or counting, we need to grow bacteria in nutrient agar plates. These are petri-dishes containing a mixture of agar and nutrients.

Peteri-dishes: Petri-dishes are disposable clear plastic dishes with a cap that are used for many science experiments and bacteria growth.  A thin layer of nutrient agar in a petri-dish is enough for growing bacteria. You can see the bacteria colony shapes and count them without opening the lead of the petri-dish. Since petri-dishes are clear, you can see the bacteria from either side of the dish.

Incubator: Incubator is a warm cabinet that you can set it's temperature to a proper temperature for bacteria growth. About 35 C is a good temperature for most bacteria. This is close the body temperature. If you be able to create such a temperature in any other way, it is as good as an incubator. You may find warm places behind your refrigerator, next to the radiator or inside an oven that is off.

You may also make an incubator by placing a small desk lamp inside a wooden or metal box. Or you may put a Styrofoam cooler upside down over a desk lamp. A small lamp (15 watts) should be able to create enough heat to warm up a small space. Prepare your incubator in advance and use a thermometer to test it a day before starting your experiment.

Material: 

  • Agar (dry powder)*
  • 10 petri-dishes (100 mmx 15mm)*
  • 1 ml Pipette*
  • 10 ml pipette*
  • 2 transfer pipette*
  • 2 test tubes with cap*
  • Glass beaker or steel pan *
  • Chicken broth or beef broth *
  • Filter Paper *

* These material are included in a kit from MiniScience.com or you may buy them individually from a local laboratory supplier.

* Chicken broth or beef broth can be purchased from supermarkets and health food stores or you may make them at home. (It must be fat free). Filter paper is coffee filter or you may substitute it with any clean cotton cloth. 

Procedure:

If you are making your own chicken broth, one small chicken can give you enough broth for this experiment. (Half a pound beef can be used instead). Boil the chicken for one hour. Separate the broth by transferring it to another pot. remove any fat from the top of the broth. Filter the broth using a clean piece of cloth or coffee filter. If you are buying chicken or beef broth from a super market, it comes in small bags of about 4 grams each. Use two bags in about 300 ml water and boil it. Then filter it and transfer the filtered broth to another clean pan.

Add water to the broth to bring it to about 650 ml and boil it again. Start adding the Agar powder while stirring.  (If you are using a MiniScience kit, entire agar tube must be used, otherwise use 8 grams of dry agar powder). You will add agar gradually while stirring. Adding agar will take 30 seconds to one minute. If you don't stir it good, agar solidifies at the bottom of the pan and burns. Continue stirring for another two minutes or until the agar solution is fully dissolved and the solution is clear. 

At this time you may optionally add a calcium carbonate tablet and some mushroom extract. (You could also boil mushroom with beef or chicken broth).

Your nutrient agar is ready at this time. Turn off the heat and let it cool down. While it is still warm set your petri-dishes on a table. Open the lead of each petri dish and pour some nutrient agar in each dish (enough to cover the bottom of the dish) and immediately put the lead back. Repeat this for all petri-dishes. Leave the dishes where they are until they solidify. 

Note: The nutrient agar should be free from all bacteria, however boiling usually is not enough to kill all the environmental bacteria that have entered our nutrient agar. For school experiments, it may not matter but in real biology laboratories, nutrient agar plates are transferred to an autoclave for sterilization. Hot, under pressure steam of autoclave can kill all bacteria in about 30 minutes. Advanced students may use a pressure cooker to do the same.

Prepare your test sample:

For counting bacteria, Use a 1 ml pipette to place 0.1 ml of your sample in a nutrient agar plate. Spread the liquid all over the plate surface using a sterile spoon. Cover the plate. Turn it upside down and place it in an incubator. Nutrient agar will remain moist when the plate is upside down. Check back within 36 hours to see the result of bacteria growth. 

When the bacteria grow, each bacteria will become a bacteria colony that can be seen as a small spot on the petri dish. 

Materials and Equipment:
Extract the list of material from the experiment section and write them here.

Results of Experiment (Observation):
Experiments are often done in series. A series of experiments can be done by changing one variable a different amount each time. A series of experiments is made up of separate experimental "runs." During each run you make a measurement of how much the variable affected the system under study. For each run, a different amount of change in the variable is used. This produces a different amount of response in the system. You measure this response, or record data, in a table for this purpose. This is considered "raw data" since it has not been processed or interpreted yet. When raw data gets processed mathematically, for example, it becomes results.
Calculations:
If you do any calculation for your project, write your calculations in this section.

Summery of Results:
Summarize what happened. This can be in the form of a table of processed numerical data, or graphs. It could also be a written statement of what occurred during experiments.

It is from calculations using recorded data that tables and graphs are made. Studying tables and graphs, we can see trends that tell us how different variables cause our observations. Based on these trends, we can draw conclusions about the system under study. These conclusions help us confirm or deny our original hypothesis. Often, mathematical equations can be made from graphs. These equations allow us to predict how a change will affect the system without the need to do additional experiments. Advanced levels of experimental science rely heavily on graphical and mathematical analysis of data. At this level, science becomes even more interesting and powerful.

Conclusion:
Using the trends in your experimental data and your experimental observations, try to answer your original questions. Is your hypothesis correct? Now is the time to pull together what happened, and assess the experiments you did.
Related Questions & Answers:
What you have learned may allow you to answer other questions. Many questions are related. Several new questions may have occurred to you while doing experiments. You may now be able to understand or verify things that you discovered when gathering information for the project. Questions lead to more questions, which lead to additional hypothesis that need to be tested.

Possible Errors:
If you did not observe anything different than what happened with your control, the variable you changed may not affect the system you are investigating. If you did not observe a consistent, reproducible trend in your series of experimental runs there may be experimental errors affecting your results. The first thing to check is how you are making your measurements. Is the measurement method questionable or unreliable? Maybe you are reading a scale incorrectly, or maybe the measuring instrument is working erratically.

If you determine that experimental errors are influencing your results, carefully rethink the design of your experiments. Review each step of the procedure to find sources of potential errors. If possible, have a scientist review the procedure with you. Sometimes the designer of an experiment can miss the obvious.

 
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