DNA Extraction from Bananas - DOC by M6E8vkJ9

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									Name:_______________________________ Period:__________ Date:__________________________
                                      DNA Extraction from Bananas

Introduction

        The process of obtaining DNA from cells is the first step in many biotechnology laboratory
procedures. Researchers must be able to separate the DNA gently from the unwanted substances in the
cells so the DNA is not broken up or sheared. The procedure for this laboratory exercise is a modified
version of the Marmur preparation. The Marmur preparation is used worldwide in biotechnology
laboratories.
        In this laboratory exercise you will make an extract of banana treated with salt, distilled water, and
detergent. The salt causes the negative phosphate ends of the DNA molecules to come closer together so
they will precipitate out of a cold alcohol solution. The detergent first disrupts the cell membranes. The
detergent then forms complexes with the lipids and proteins released from the cell, causing them to
precipitate out of solution. Finally, cold alcohol will be added to the extract to separate out the DNA.

I. Materials

a) Beaker of Water                            h) Plastic funnel
b) Beaker to hold alcohol                     i) Glass stirring rod
c) 1 – 25 mL graduated cylinder               j) Coffee filter or thin filter paper
d) 1 – 10 mL graduated cylinder               k) Extraction buffer (a salt solution)
e) Clean beaker                               l) Detergent
f) Clean test tubes                           m) Banana
g) Petri dish                                 n) Fork

II. Procedure

1. Squash a small piece of banana (2-3 cm) with a fork in a Petri dish.

2. Measure out 12 mL of the extraction buffer and put it into the Petri dish with the banana.

3. Measure out 3 mL of detergent and add it to the banana solution in the Petri dish to break open the
   cells. Mix well using a stirring rod, but not enough to make bubbles. The detergent dissolves the
   lipids that hold the cell membranes together – this releases the DNA into the solution. The detergent
   further causes the lipids and proteins to precipitate out of the solution, separating them from the
   DNA.

4. Dampen (with water) a coffee filter or thin filter paper (fold the filter into a cup-like shape) and
   place it in a funnel. Put the funnel into a clean beaker and pour the banana mixture into the funnel.
   Collect the filtrate into the glass vial. Throw away the filter paper and banana remains when finished.

5. Measure out 1 mL of the filtered banana juice and put it in a glass test tube.

6. Add 1 mL of water and mix well.

7. Carefully and slowly pour 8 mL of cold alcohol down the side of the tilted test tube so that the alcohol
   remains in a layer above the juice. Because the alcohol has a lower density, the alcohol will “float” on
   top of the juicy solution.
Name:_______________________________ Period:__________ Date:__________________________
8. Let the solution sit for 2-3 minutes. You will see DNA precipitate out into the alcohol layer. A white
   substance will become visible at the interface where the two liquids meet – this is DNA! DNA is clear
   or may appear kind of white. You may have a tough time seeing it at first. Bubbles will form on the
   DNA strands, and this should help you to recognize it.

The alcohol will form a layer on top of the cell scum. The cell scum is heavy and will remain on the
bottom of the vial. The DNA is less dense than the cell scum, but more dense than the alcohol. Thus, the
DNA will stay in the middle, where the two layers meet. Since DNA cannot remain dissolved at the
alcohol concentration used here, it begins to come out of solution or precipitate. When it does, the DNA
will start to rise up into the alcohol layer.

Optional Activity: Perform this step carefully because this will sometimes cause your DNA to break up
into small pieces. You may use a glass rod to slowly swirl the DNA in the alcohol layer (not the lower
layer!). You will begin to be able to SEE the DNA. See if you can swirl enough DNA onto your rod to
pull it out of the test tube.

III. Analysis

1. What is the purpose of the salt solution in the beginning of the procedure?




2. What is the purpose of the detergent during the extraction process?




3. Why do you think alcohol is used in this experiment to precipitate DNA and not a different solution?




4. Describe the characteristics of the extracted DNA, such as color, shape, size, and consistency.




5. Where is the DNA found in the cell?




6. Why will scientists use the Marmur preparation procedure?
Name:_______________________________ Period:__________ Date:__________________________
IV. Post-lab/ DNA Practice Questions

1. What does DNA stand for?


2. Who were the two scientists that discovered the structure of DNA and created the first model of DNA?


3. What category of organic molecule (carbohydrate, lipid, protein, or nucleic acid) is DNA?


4. What is the monomer of DNA?


5. What are the three parts of a DNA nucleotide?


6. Why is DNA said to be a double helix?



7. What are the four bases that make up the rungs of DNA?



8. What kinds of bonds hold the bases together to create the rungs of DNA?


9. What is Chargaff’s Rule?



10. Who took pictures of DNA?


11. Given the following sequence of bases, what are the complementary bases (remember Chargaff’s
    rule!!!)?

       A   T G     C    C     C   T   A T   A G      C   G   C   G    C   A   A   A G      A G

								
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