Awesome After School Kits
Isolation of Fruit DNA
Objective:
Youth will:
- Perform simple DNA
isolation while learning
basic lab skills.
- Explore the nature of
DNA while isolating the
DNA from different
biological materials.
Grade Level: 5th-8th
Academic Standards:
- Youth will learn the
importance of following
proper laboratory
procedures.
- Observing, comparing,
organizing, relating
Life Skills:
- Learning to learn,
communicating
Time: 30-45 minutes
Supplies Needed: See
list
Advance Preparation:
Gather material for DNA
isolation.
Notes: Alcohol or
isopropyl should be used
in a well-ventilated area.
Curriculum to Use: University of Kentucky, College of Agriculture, Plant & Soil Science Department. Dr. David Hildebrand, Scott Serdoz. Adapted by University of Illinois Extension from Iowa State University Extension and Iowa State University Office of Biotechnology Presenter Information: DNA is discussed as being sub‐microscopic, but if you get enough of it together in one place in the right environment, it becomes visible. Actually seeing the mass of DNA created as the end result of these activities brings home the lesson that DNA is real and in every living organism. In general, the process of DNA extraction from the nucleus of a cell requires the following: 1. Breaking cell and nuclear membranes mechanically (blending, mixing) and chemically (with detergents). 2. Separation of cellular debris from DNA (filtration—optional in some procedures). 3. Precipitation and clumping of thousands of DNA molecules using alcohol. These activities are similar to the work that is practiced in biotechnology and genetics laboratories around the world. DNA extraction is also the first step in genetic engineering. Fruits or onions with low starch content are used because they allow the DNA to be seen clearly. The salt shields the negative phosphate ends of DNA, which allows the ends to come closer so the DNA can precipitate out of a cold alcohol solution. The detergent causes the cell membrane to break down by dissolving the lipids and proteins of the cell and disrupting the bonds that hold the cell membrane together. The detergent then forms complexes with these lipids and proteins, causing them to precipitate out of the solution. Activity Introduction: The cells of all living organisms contain DNA. Extracting DNA is the first step in many biotechnology laboratory procedures. Scientists attempt to separate the unbroken DNA strands from any unwanted substances. This technique is important to any scientist working with DNA, whether making recombinant DNA or mapping the human genome. This procedure is designed to extract DNA from fruit or vegetables in sufficient quantity to be seen and spooled. It is based on the use of household equipment and supplies. These activities are similar to the work that is practiced in biotechnology and genetics laboratories around the world. DNA extraction is also the first step in genetic engineering. Awesome After School Kits
Materials: ‐ Fruit (bananas, strawberries, etc.) ‐ Salt ‐ Baking soda ‐ Dish detergent (liquid) (Dawn works well) ‐ Rubbing alcohol Note: alcohol/isopropyl should be used in a well­ventilated area. ‐ Coffee filters or porous, non‐absorbent cloth (i.e. bed sheet pieces) ‐ Large paper clips or tooth picks ‐ Narrow bottle, jar or test tube; small spoon or mortar and pestle ‐ Water Activity: 1. “DNA extraction solution”: add 1 tsp baking soda, 1 tsp detergent and ¼ tsp salt to ½ cup of water in a mug, bottle. 2. Put approximately ½ inch slice of fruit in a small bowl and smash with a spoon or mortar and pestle. 3. Add 3‐4 tsp “DNA extraction solution” to the bowl and grind (extract) with the spoon until the fruit is well pulverized. 4. Place the cloth or filter on top of the narrow bottle, jar or test tube and push part way in. Filter the DNA extract. 5. Add an equal volume of the cold rubbing alcohol, mix by swirling. Wait approximately 5 minutes or more. 6. Straighten large paper clip and make a small hook (~ 3 mm) at one end to “catch” the DNA. “Fish” out precipitated DNA with paper clip hook. When extracting the DNA, twist the toothpick or paper clip slowly. Be sure to only remove the DNA from the top layer. Extract the DNA that floats to the surface. Talk It Over: After explaining that they’re going to extract DNA from fruit, ask your students the following questions: What does it mean to extract? To extract means to draw out by effort, to take out or to remove. Where is DNA found? In the nuclei of plant and animal cells, every living cell contains DNA. Why would we want to extract DNA from fruit? Soft fruits work best for this experiment because the cell membranes are easily broken down. Why would scientists want to extract DNA from an organism? Extracting DNA is the first step in many biotechnology laboratory procedures used to study or change the DNA. Sharing ‐ • By mashing the banana you are physically disrupting the cells and cell membranes are broken open, allowing the DNA to escape. • Mixing the banana with the extraction solution allows the lipids and proteins to be solublized by the detergent. • The baking soda in the solution acts as a buffer to keep the solution at an optimal ph for DNA (approx. ph=7). • When poured through the coffee filter the cellular debris is trapped, however, the DNA is able to flow through. • The salt, in combination with the alcohol (isopropyl), precipitates the DNA, because the DNA is not soluble in alcohol (isopropyl). This allows the DNA to be seen by the naked eye. DNA has a negative charge and the salt acts to neutralize this charge so the DNA can remain in its helix form. 2
Awesome After School Kits
Processing – Where you able to extract some DNA? ‐ What part of the procedure do you think released the DNA? DNA is released by agitation and by the action of the detergent. ‐In what step did you separate the DNA from the cellular debris? By filtration Generalizing – How are the jobs of detectives and scientists similar? Both scientists and investigators use logical procedures to find solutions to problems. Applying – Extend learning beyond this lesson. Different media for Lab stations: You may use a variety of fruits or vegetable media in this DNA extraction lab procedure. Onion, kiwi, lettuce and broccoli all have been successful in producing good results in the extraction procedure. The main requirement is that the youth must slice up or cut the media into small pieces to facilitate the breaking down of the cell structure, allowing for a good production of DNA into solution. Bananas or Strawberries are the easiest to use. You may choose two or three media for the groups to use so they can compare the production of DNA from each. You will need one onion, a quarter head of lettuce, two kiwi, or one third of a bunch of broccoli, 1 banana per group, or several strawberries for each extraction station you set up. It provides a strong point of discussion leading to the fact that DNA in plants, animals and other living organisms are all made up of the same building block proteins. The youth will not be able to distinguish between kiwi and onion DNA or in any other comparison of extracted DNA. Presenter Tip: Use different fruits, vegetables for comparison stations 1. When pouring the alcohol, make sure that two separate layers are being formed (The bottom layer being the banana mixture and the top layer being the alcohol). 2. When extracting the DNA, twist the toothpick or paper clip slowly. Be sure to only remove the DNA from the top layer. ‐ This activity can be done at science fair nights, county fairs and short term science introductions. Sources: [any additional curriculum, websites, books, etc. that were or could be used with this lesson.] Extended Learning: Blender protocol (http://biology.about.com/c/ht/00/07/How_Extract_DNA_Banana0962932481.htm) 1. Gather together a banana, salt, warm water, a blender, liquid soap, toothpicks or paper clips, a strainer, a glass jar and rubbing alcohol. 2. Cut your banana into small pieces. 3. Place your banana pieces in the blender, add a teaspoon of salt and slightly cover the mixture with warm water. 4. Mix in the blender for 5 to 10 seconds making sure the mixture is not too runny. 5. Pour the mixture into the glass jar through the strainer. You want the jar to be about half full. 6. Add about 2 teaspoons of liquid soap and gently stir the mixture. You should try not to create bubbles when stirring. 7. Carefully pour the rubbing alcohol down the side of the glass stopping near the top. 8. Wait for 5 minutes. 9. Use the toothpicks or bent out paper clips to extract the DNA that floats to the surface. Adapted by University of Illinois Extension from Iowa State University Extension and Iowa State
University Office of Biotechnology and University of Kentucky,
Plant & Soil Science Department.
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