DNA Extraction
An organism’s genetic information is encoded in its DNA, or deoxyribonucleic acid. DNA is
assembled from sequences of nucleotides, each of which contains a deoxyribose sugar, a phosphate
group, and one of the four bases: adenine, guanine, cytosine, or thymine. Specific regions of DNA are
called genes, with each gene containing the information for the assembly of a particular protein. DNA,
when found in cells, is usually packaged with proteins bound to it. When we look at cells under the
microscope, we see a large prominent organelle called the nucleus where the DNA is located. When we
visualize this under a microscope we see this DNA-protein complex and refer to it as chromatin. During
certain phases of the cell cycle the chromatin is very tightly coiled and wound into visible structures
called chromosomes. A lot of DNA is packed into each cell. If we could stretch out all he DNA into one
long string, it would be more than six feet long! So, you can imagine that the DNA molecule is very thin.
ISOLATION OF DNA
Isolation of DNA from cells is a routine procedure used by molecular biologists. Once isolated
the DNA can be characterized and manipulated in a variety of ways. When DNA is expelled and isolated
from cells, the solution in which it is found is very viscous. This viscosity is due to the fact that DNA
molecules are very long and thin and tend to stick to each other. Remember hydrogen bonds?
Hydrogen bonds not only hold the double helix together at the base pairs, but hydrogen bonds among
the DNA molecules make them tend to stick together. If we disrupt these hydrogen bonds with harsh
chemicals or heat we tear the DNA molecules apart. How does this effect viscosity?
In lab today we will extract DNA from strawberries, and then work with the extracted DNA to
see how chemicals or heat affect viscosity. But first, we will take a closer look at the structure of DNA.
DNA STRUCTURE
Nucleic acids are long, chainlike molecules formed by linking together large numbers of
nucleotides. DNA and RNA are nucleic acids. What are the three main differences between DNA and
RNA?
When DNA is extracted from multiple cells, the amassed quantity is visible and looks like strands of
mucous-like, translucent cotton.
In class, we created DNA keychains using beads to depict phosphate groups, deoxyribose (sugar) and our
4 nitrogenous bases. Remember what DNA looks like as we extract it from our strawberry.
DNA Extraction
Materials Per Person
1-15 mL tube
10 mL aliquot of DNA extraction buffer
5-mL aliquot of 95% ethanol in transfer vessel
1 strawberry
1 50mL tube
1 piece of cheesecloth
1 funnel
1 wooden stick
resealable bag
1 transfer pipet
Methods
1. Obtain one fresh or frozen and thawed strawberry. If you are using a fresh strawberry, remove
the green sepals (tops) from the berry.
2. Place the strawberry in a resealable bag.
3. Close the bag slowly, pushing all of the air out of the bag as you seal it.
4. Being careful not to break the bag, thoroughly mash the strawberry with your hands for two
minutes.
5. Pour the 10 mL aliquot of extraction buffer into the bag with the mashed strawberry. Reseal the
bag.
6. Mash the strawberry for one additional minute.
7. Place a funnel into a 50 mL centrifuge tube. Fold the cheesecloth in half along the longer edge
and place into the funnel to create a filter. The cheesecloth will overlap the edge of the funnel.
8. Pour the strawberry mixture into the funnel, filtering the contents through the cheesecloth and
into the 50 mL centrifuge tube.
9. Carefully pour 2 mL of the filtered contents from the 50 mL tube into a clean 15 mL tube. Use
the lines on the side of the 15 mL tube to help measure the amount added.
10. Hold the 15 mL tube at an angle. Using a transfer pipet, carefully add 5 mL of cold 95% ethanol
by running it down the inside of the tube. Add the 95% ethanol until the total volume is 7 mL
(use the lines on the side of the tube to help you measure). You should have two distinct layers.
Caution: Do not mix the strawberry extract and the ethanol.
11. Watch closely as translucent strands of DNA begin to clump together where the ethanol layer
meets the strawberry extract layer. Tiny bubbles in the ethanol layer will appear where the DNA
precipitates.
12. Students should work as pairs from this point. Use one of the test tubes and slowly and
carefully rotate the wooden stick in the ethanol directly above the extract layer to wind (or
spool) the DNA. Remove the wooden stick from the tube and observe. (Write your observations
down in your lab notebook).
Part II
13. Now, take the other test tube you prepared to investigate how heat, or a change in pH can
affect DNA. Some groups will work on the affects of heat while other groups work on the affects
of pH. Decide which groups will do which experiment, then proceed.
EFFECTS OF HEAT
1. Use the second tube of extract.
2. Place the tube into a boiling water bath for 10 minutes.
3. Remove, and place the tube into an ice bath
4. Insert and twirl a wooden stick into the tube and spool the DNA.
5. Compare the viscosity of the heat treated DNA with the untreated DNA from your first tube.
What effect does heat have on the viscosity of DNA?
What do you think is the mechanism for this change in viscosity?
EFFECTS OF pH ON DNA
1. Use the second tube of extract.
2. Add 2 ml of 1.0 N NaOH to the tube
a. NaOH is a caustic base. Be careful not to spill on yourself, your clothes, or the benches
b. Notify the instructor of any spills.
3. Insert and twirl a wooden stick into the tube and spool the DNA.
4. Compare the viscosity of the alkali-treated DNA with the untreated DNA from your first tube.
What effect does a basic pH have on the viscosity of DNA?
What do you think is the mechanism for this pH-induced change in viscosity?