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Lab-DNA Extraction

DNA Extraction Lab

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arreolakaren6801
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11 views

Lab-DNA Extraction

DNA Extraction Lab

Uploaded by

arreolakaren6801
Copyright
© © All Rights Reserved
Available Formats
Download as DOCX, PDF, TXT or read online on Scribd
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Lab: DNA Extraction

(Based on the work of the Office of Biotechnology, Iowa State University)

Introduction

DNA is present in the cells of all living organisms. This procedure is not unlike the procedure biotechnology
companies use to extract DNA from the cells with which they are working. Like biotechnology companies, we
use detergent and salt to break down cell structures and release the DNA into a solution. Biotech firms use
centrifuges to isolate the DNA from other molecules by its density. Instead of a centrifuge, this procedure
uses alcohol to “float” or precipitate the DNA out of the water/detergent/salt solution.

Denaturation is the alteration of a protein shape through some form of external stress (for example, by
applying heat, acid or alkali), in such a way that it will no longer be able to carry out its cellular function.

A buffer is an aqueous solution consisting of a mixture of a weak acid and its conjugate base or a weak
base and its conjugate acid. Its pH changes very little when a small amount of strong acid or base is added to it
and thus it is used to prevent changes in the pH of a solution. Buffer solutions are used as a means of keeping
pH at a nearly constant value in a wide variety of chemical applications. Many life forms thrive only in a
relatively small pH range so they utilize a buffer solution to maintain a constant pH. One example of a buffer
solution found in nature is blood

Objectives:
● Know how to extract DNA from fruit

● Observe what DNA looks like from the naked eye

● Understand that DNA is in all living things and once living things

● Understand the terms: DNA, denature, extraction

Materials (per group)

electronic balance
weigh boats
2 x 150 mL beaker
50 mL beaker
1 g Salt
45 mL distilled water
5 mL Dawn dishwashing detergent
30g peeled kiwi fruit
Fork
sheet of waxed paper
coffee filter
test tube
5 mL 99% isopropyl alcohol
4 inches of thin wire with hooked end, or inoculation loop
thermometer
hot water bath (50-60 degrees C)
ice water bath
Procedures

1. (2:00pm)Dissolve 1 g salt in 45 mL distilled water in a 150 mL beaker


2. (2:02)Add 5 mL of Dawn detergent and stir it gently to prevent foaming
3. Peel the fruit over waxed paper and cut into chunks with mass approximately 30 g
4. (2:03)Mash 30 g of kiwi fruit on the waxed paper with a fork and place the mashed fruit in a 150 mL
beaker
5. (2:04)Pour the detergent/salt solution over the fruit so that the fruit is JUST COVERED with the solution,
then stir gently to mix. (You may not need to use all of the solution you have prepared.)
6. (2:24)(Place the 150 mL beaker with the solution and the fruit in the hot water bath that measures 60o C
for 15 minutes. During this time, mash the fruit against the side of the beaker every three to four
minutes.
7. After 15 minutes, immediately place the 150 mL beaker with the solution and the fruit into the ice water
bath for 5 minutes. Continue to mash the fruit with the fork as it gradually cools.
8. (2:26)Wash out the 150 mL beaker.
9. (2:27)Seat the coffee filter over the 150 mL beaker.
10. (2:30)Pour the fruit solution into the filter and strain until most of the liquid (filtrate) is recovered.
11. (2:39) Swirl the filtrate in the beaker before pouring it into a test tube. (This is very important, as solids
that may contain DNA can accumulate at the bottom of the beaker.)
12. (Pour 5 mL of filtered fruit solution into a test tube.

13. Pour 5 mL of 99% isopropyl alcohol on top of the fruit filtrate, forming 2 layers. Tip the test tube and try
to pour the alcohol along the side of the test tube so that the filtrate is disturbed as little as possible.
14. Make a hook at the end of the wire and hook some of the DNA that has precipitated out of the solution.
15. Place a small amount of DNA on a microscope slide, cover with a cover slip and observe under the
microscope at high power. Adjust the light diaphragm to see the DNA strands better

Data:
1. Record your initial observations of the DNA sample you have collected.

There are little white stuff floating in the alcohol.

2. Record your observations of the DNA sample you have collected under the microscope.
Conclusion:
1. What was the purpose of mashing up the fruit?
To get the most of the DNA from the fruit.

2. What does the extraction buffer do? (Hint: Extraction buffer contains soap. What does soap do
when you wash your hands?)
Soap cleans the germs that might interfere with looking clearly at the DNA.

3. What does the filter do?


The filter extracts all the liquid from the kiwi.

4. What happened when you added the filtrate to the alcohol?


The alcohol floats on top of the liquid (doesn’t mix).

5. What did the DNA look like without magnification? With magnification?
You cannot see DNA with the naked eye but with magnification you can see a double helix within
a chromosome.

6. Is DNA found in all living or once living cells?


DNA is found in all living cells.

7. In the lab what was used to denature the fruit’s cells?

8. What are the parts of the DNA?


DNA is made up of three things: deoxyribose (a sugar), a phosphate group, and nitrogenous
bases.

9. Given a sequence of a DNA strand as a template, write the complementary strand for:
TAC ATC GCG GTA ACG ATC GTT ATA GCA ACT
ATG TAG CGC CAT TGC TAG CAA TAT CGT TGA

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