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DNA Purification by Precipitation
Method
1. Weigh out 10 g of the chopped onion in the weighing boat. (Chopping breaks the cells
apart from one another and provides a large surface area over which to extract the DNA.).
Place into the mortar.
2. Add 1 cm3 of detergent, one spatula of salt and 10 cm3 water. Use the pestle to mash all
the ingredients together. Don't stir too fast or else you'll get lots of bubbles!
3. Place this mixture into the 100ml beaker, write your initials on the beaker and incubate at
60oC for 15 minutes. Start the stop clock. Incubation helps to break up the cells further
and starts to degrade some of the cell's proteins. Whilst you are waiting, start answering
the questions on this sheet.
4. Add two to three drops of protease enzyme to the boiling tube.
5. Remove the beaker from the incubator and filter the onion mixture through a fine sieve
into the boiling tube. This removes all the unwanted lumps and bits of onion. You should be
left with a green liquid, and this contains the onion DNA.
6. Take 4 cm3 (two pipettes) ice cold alcohol and pour it slowly down the side of the boiling
tube using the plastic pipette. Don’t do this drop-by-drop). The alcohol will form a
transparent layer on top of the onion mixture, as the alcohol is less dense.
7. Where the layer of ice cold alcohol meets the onion mixture
underneath, you will see a white jelly-like substance forming.
This is the DNA. You can remove the DNA using the blue hook
DNA
in your tray.
Main points:-
· Macerate tissue
· Detergent
· Alcohol to precipitate the DNA
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Questions:
1. Here is a diagram of the structure of DNA.
a. Draw and label one monomer of DNA in
the space below. (3 marks)
b. What is the name of the bonds holding the two strands of
DNA together? (1 mark)
c. Complete the table to show the names and percentages of
the other bases. (5 marks)
Nitrogenous base Percentage of DNA (%)
Adenine 36
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2. DNA is a nucleic acid. Describe the structure of DNA, using the following key words: purine,
pyrimidine, hydrogen bonds, phosphodiester bond, double-helix, complementary base pairing,
sugar-phosphate backbone, nitrogenous bases. Underline the keywords in your answer.
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3. In eukaryotic cells, DNA is associated with proteins.
a. What are these proteins called?
b. Describe how these proteins are associated with DNA. You may draw a diagram to help
you.
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4. When purifying DNA by precipitation, various tools are used to help extract the DNA from the
nuclei of plant cells. Suggest the role of each of the following:
a. The pestle and mortar
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b. The detergent
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c. The salt solution
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d. The water bath
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e. The protease enzyme
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5. During the process of extracting and purifying DNA, the hydrogen bonds
between the nitrogenous bases are broken. The diagram shows a section of a
strand of DNA.
In the space below, draw the complementary DNA bases onto the strand,
showing the number of hydrogen bonds between the bases. (2 marks)
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Stretch and challenge questions (A Grade):
6. Suggest why the DNA precipitates out of the solution.
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7. Suggest a method that could be used to prove that the white substance is DNA.
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Very challenging question (A* Grade)
8. Read about semi-conservative replication below, then answer exam question 32 from your exam
question booklet.
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