The purification of DNA 2.1.3 dii

Question 1

Why are polynucleotides anti-parallel?

Answer 1

Due to the orientation of the sugar-phosphate backbone, meaning one of them runs from the 5’-3’ direction and the other runs in the 3’-5’ direction.

Question 2

How is DNA purified using precipitation?

Answer 2

1. Break up the cells
2. Make up a detergent solution (washing up liquid, salt and water)
3. Add cells to detergent and leave for 15 mins at 60 degrees in a water bath
4. Cool mixture in ice bath, then filter and transfer to a clean boiling tube
5. Add protease enzymes to break down the proteins bound to DNA
6. Dribble cold ethanol down the side of the tube
7. Leave for a few minutes
8. Observe white precipitate and remove using a glass rod = DNA

Question 3

How does DNA extraction work in a kiwi fruit?

Answer 3

The salt and detergent mix attacks the cell membranes of the kiwi fruit - releasing all the cell contents out (nucleus etc.).
DNA within the nucleus is surrounded by proteins; which are broken down by the enzyme proteinase (found in a kiwi fruit) which frees the DNA.
The DNA then turns solid when the methylated spirits are added - forming a white precipitate.

Question 4

Explain the purpose of a detergent in DNA extraction

Answer 4

Breaks down the cell membrane

Question 5

Quick question - The precipitate contains a mixture of DNA and RNA. Suggest what could be done to obtain a pure sample of DNA

Answer 5

Add an enzyme RNase to break down RNA before adding the ethanol