Download An Apple a Day: Extracting DNA from Any Living Thing

An Apple a Day:
Extracting DNA from Any Living Thing
EXTENDED VERSION
Unit:
The Apple Genomics Project
Lesson:
An Apple a Day: Extracting DNA from Any Living Thing
Audience:
This lesson is intended for use with middle and high school science students.
Student Learning Objectives:
At the completion of this lesson the students will be able to:
1.
2.
3.
4.
5.
6.
7.
Define the process of DNA extraction.
Explain the purpose of the extraction process.
Explain why DNA is essential and a part of all living things.
Explain where DNA is found.
Demonstrate the ability to follow instructions in a lab experiment.
Explain the function of each material in a lab experiment.
Compare results of the exercise using different sources for DNA extraction.
Student Activities:
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Take notes on lesson/pre-lab content as presented by instructor.
Complete pre-laboratory worksheet.
Complete laboratory exercise.
Discuss laboratory exercise.
Complete post-laboratory worksheet.
Problems & Questions for Study:
1.
2.
3.
4.
5.
6.
7.
Where is DNA found?
Why is DNA essential to all living things?
What is DNA extraction?
What is the purpose of the extraction process?
What are the steps in the DNA extraction process?
What is the function of each material in this lab experiment?
How do the results of the exercise change, using different sources for DNA
extraction?
Documentation of Competencies and Academic Standards Met:
National Academic Standards
Science Content Standard A for Grades 9-12: Develop abilities necessary to do scientific
inquiry.
Science Content Standard B for Grades 9-12: Develop understanding of chemical
reactions.
Science Content Standard C for Grades 9-12: Develop understanding of the cell.
Connection to Supervised Agricultural Experience/Career Development
Careers related to horticulture include fruit grower, extension agent, plant cytologist,
flower grader, bacteriologist, grounds keeper, and tree surgeon. [For more careers, visit
the FFA Career Center at
http://www.ffa.org/collegiate.cfm?method=c_job.CareerSearch.]
FFA proficiency areas related to horticulture include floriculture, landscape architecture,
and specialty crop production. [For more proficiency areas, visit the National FFA
website for proficiencies at http://www.ffa.org/programs/proficiency/index.html.]
Connection to FFA/Leadership Development/Personal Growth
Career Development Events (CDEs) related to horticulture include floriculture and
nursery & landscaping. [For more information on CDEs, visit
http://www.ffa.org/programs/cde/index.html.]
Motivation/Interest Approach:
Display various items to the class, including “living”, “never living”, and “previously living” objects.
Ask students to explain what DNA is. Once the class comes up with a definition of DNA, ask
students to identify which objects contain DNA. Have students sort the objects into the three
categories (living, never living, or previously living) based on their identifications. Ask students to
explain how and why they categorized each object the way they did. Most likely, students will sort
objects into only two categories: “living” and “non-living”. If this is the case, point out that some of
the objects were “previously living”. Ask students whether these objects have DNA, since they were
alive at a previous time. Initiate a discussion in which students may re-categorize objects, if
necessary, to include “living”, “never living”, and “previously living” objects. Ask students how they
know that “living” cells (and “previously living” cells) contain DNA, if they cannot see physical
proof of the DNA itself. Explain to students that the laboratory exercise in this lesson will allow them
to extract and observe DNA from living cells.
Content Outline & Teaching Procedures:
I. Background
A. DNA ~ deoxyribonucleic acid
B. DNA is present in EVERY cell of ALL living organisms.
C. Long, thick fibers, arranged in a double helix, of DNA store information for the
functioning of the chemistry of life, and for building and maintaining a living organism.
D. The process of extracting (taking it out of, or removing it from) DNA from a cell is the
first step for many laboratory procedures in biotechnology, including gel electrophoresis and
DNA sequencing.
1. Scientists use DNA extraction to separate DNA from the unwanted substances of
the cell, in order to examine the DNA.
2. It must be done gently enough so that the DNA does not denature (break up).
II. Laboratory Exercise
A. DNA is the “blueprint for life,” and it can be extracted from any living (or formerly
living) thing.
1. Examples of things with DNA that we could extract: Split peas, spinach, chicken
liver, onions, and broccoli. (We’ll use bananas for this exercise, because they have a
LOT of DNA packed into cells that are easy to break open.) All of these are things
that are living.
2. Examples of things with DNA that we could extract, but probably would not want
to try extracting: Wood, preserved fossils, cotton. These are things that are living, or
were previously living, but DNA extraction would be difficult.
3. Examples of things with no DNA: Rocks, water, candles, plastic, and pop cans. All
of these are things that never lived.
B. DNA found in banana cells can be extracted using common materials. This procedure uses
household equipment and store supplies to extract DNA from bananas in sufficient quantity
to be observed and spooled.
1. In this exercise, you will prepare a solution of banana treated with salt, distilled or
tap water, and Dawn dishwashing detergent (or Suave clarifying shampoo).
a. Salt breaks up the protein chains that bind the nucleic acids. The salt allows
the DNA to precipitate out of a cold alcohol solution.
b. Dawn dish soap is a detergent, so it dissolves the lipid (fat) part of the
banana cell wall and nuclear membrane, allowing the DNA inside to spill out.
i. The detergent breaks down the cell membrane by dissolving the
lipids (fatty molecules) and proteins of the cell and disrupts the bonds
that hold the cell membrane together.
ii. The detergent then forms complexes with these lipids and proteins,
which are filtered out of solution by the coffee filter, while leaving
the cells' DNA in the filtrate (the liquid solution that goes through the
coffee filter into the cup).
c. A blender will be used to separate the banana cells from each other,
resulting in a thin banana soup.
2. Successful spooling will result in clumps of white, stringy DNA.
a. The white, stringy stuff is actually a mix of DNA and RNA.
b. The procedure for DNA extraction is really a procedure for nucleic acid
extraction. Much of the RNA is cut by ribonucleases (enzymes that cut RNA)
that are released when the cells are broken open.
C. Troubleshooting the laboratory exercise
1. Problem: No DNA precipitates out of banana/detergent/alcohol solution.
a. Solution 1: First, check one more time for DNA. Look very closely at the
alcohol layer for tiny bubbles; clumps of DNA are often loosely attached to
the bubbles.
b. Solution 2: If you still do not see DNA, then you need to make sure that
you started with enough DNA in the first place. Many food sources of DNA,
such as grapes, also contain a lot of water. If the blended cell soup is too
watery, there will not be enough concentrated DNA to see. To fix this, go
back to the first step and add less water. The cell soup should be opaque (you
should not be able to see through it).
c. Solution 3: If you still do not see DNA, then you need to make sure that
you allow enough time to pass for each step. Make sure to stir in the detergent
for at least five minutes. If the cell and nuclear membranes are still intact, the
DNA will be stuck in the bottom layer. Often, if you let the
banana/detergent/alcohol mixture sit for 30-60 minutes, DNA will precipitate
into the alcohol layer.
2. Problem: DNA clumps together.
a. Solution: None. Single molecules of DNA are long and stringy. For
example, each cell of your body contains six feet of DNA, but it's only onemillionth of an inch wide. To fit all of this DNA into your cells, it needs to be
packed efficiently, so DNA twists tightly and clumps together inside cells.
Even when you extract DNA from cells, it still clumps together, though not as
much as it would inside the cell.
D. Using this DNA as a sample for gel electrophoresis.
1. You can try this, but all you will most likely see is a smear.
2. The DNA you have extracted is genomic, meaning that you have the entire
collection of DNA from each cell. Unless you cut the DNA with restriction enzymes,
it is too long and stringy to move through the pores of the gel.
E. Other things to have students try:
1. Experiment with other DNA sources. Which source gives you the most DNA?
Why do you think this is the case? How can you compare them?
2. Experiment with different soaps and detergents. Try powdered soaps, clear
shampoo, body scrub, meat tenderizer, pineapple juice, or contact lens cleaning
solution.
3. Try leaving out a step or changing how much of one ingredient you use.
References & Teaching Aides:
Websites:
•
DNA in My Food? The Making of a Smoothie:
http://www.biotech.iastate.edu/publications/lab_protocols/DNA_Extraction_Smo
othie.html
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The Science Behind Our Food: Lesson in a Trunk: Banana DNA Extraction:
http://devacaf.caes.uga.edu/main/index.cfm?page=liatpics&trunkName=Banana%
20DNA%20Extraction
•
Genetic Science Learning Center at the Eccles Institute of Human Genetics at the
University of Utah: http://gslc.genetics.utah.edu/units/activities/extraction/
Books:
•
DNA Is Here to Stay, by Dr. Fran Balkwill. 1992. ISBN # 0-00-191165-1. (Ages
9-15)
•
Amazing Schemes within Your Genes, by Dr. Fran Balkwill. 1994. ISBN #
0876146353. (Ages 9-12)
•
Double-Talking Helix Blues, by Joel Herskowitz. 1993. Book and cassette tape.
ISBN # 0-87969-431-9. (Ages 8 and up)
•
Ingenious Genes, by Patrick A. Baeuerle and Norbert Landa. 1998. Barron's
Educational Series, Inc., Hauppauge, NY. ISBN # 0-7641-5063-4. (Ages 9-12)
•
How the Y Makes the Guy, by Norbert Landa and Patrick A. Baeuerle. 1998.
Barron's Educational Series, Inc., Hauppauge, NY. ISBN # 0-7641-5064-2. (Ages
8-12)
Additional Materials:
Handouts
An Apple a Day: Class Notes
Worksheets
An Apple a Day: Pre-Laboratory Worksheet
An Apple a Day: Post-Laboratory Worksheet
Laboratory Worksheets
An Apple a Day: Laboratory Exercise
An Apple a Day: Laboratory Exercise, Part 2
Name: ___________________________________
An Apple a Day: Class Notes
I. Background Information
A. DNA is the acronym for ______________________________ ________________.
B. DNA is present in _________________ cell of ________________ living organisms.
C. Long, thick fibers of DNA store _________________ for _______________________
______________________________________________, and for __________________
_______________________________________________________________________.
D. The process of extracting DNA from a cell is the first step for ___________________
_________________________________, __________________________________ and
__________________________________.
1. Scientists separate _________________ from the _______________________
________________________________________________________, in order to
___________________________________________________.
2. This must be done gently enough so that _______________________________
_________________________________________________________________.
II. Information about the Laboratory Exercise
A. Since DNA is the “__________________________________,” it can be extracted
from __________________________________.
1. Examples of things with DNA that we could extract: _____________________
_________________________________________________________________.
(We’ll use _________________ for this exercise, because they have a LOT of
DNA packed into cells that are easy to break open.) All of these are things that are
_________________.
2. Examples of things with DNA that we could extract, but probably would not
want to try extracting: _______________________________________________
__________________________________________________. All of these are
things that are _________________, or were _____________________________.
3. Examples of things with no DNA: ____________________________________
________________________________________. All of these are things that
__________________________________.
B. The DNA found in banana cells can be extracted using _________________________
______________________.
This procedure uses _______________________________________________________
and __________________________________ to extract DNA from bananas in sufficient
quantity to be _________________ and _________________.
1. In this exercise, you will prepare a solution of _________________ treated
with _________________, _________________, and _________________.
a. Purpose of salt: _____________________________________________
____________________________________________________________
___________________________________________________________.
b. Dawn dish soap is a type of _________________. Purpose of Dawn
dish soap: ___________________________________________________
____________________________________________________________
___________________________________________________________.
i. The detergent ______________________________________ by
_____________________________________________ of the cell
and __________________________________ that hold the cell
membrane together.
ii. The detergent then forms ___________________________ with
_____________________________________, allowing them to be
________________________________________________ by the
_________________, while leaving the cells' _____________ in
the filtrate.
c. We’ll use a __________________ to mix the banana solution. This step
will ______________________________________________, resulting in
___________________________________________________________.
2. Successful spooling will result in ____________________________________.
a. The white, stringy stuff is actually a mixture of _______ and _______.
b. The procedure for DNA extraction is really a procedure for __________
_____________________________________. Much of the RNA is cut by
____________________ (enzymes that cut RNA) that are released when
the cells are ____________________.
B. Troubleshooting the laboratory exercise
1. Problem: No DNA precipitates out of banana/detergent/alcohol solution.
a. Solution 1: ________________________________________________
____________________________________________________________
____________________________________________________________
___________________________________________________________.
b. Solution 2: ________________________________________________
____________________________________________________________
____________________________________________________________
___________________________________________________________.
c. Solution 3: ________________________________________________
____________________________________________________________
____________________________________________________________
___________________________________________________________.
2. Problem: DNA clumps together.
a. Solution: ________________________________________________
____________________________________________________________
____________________________________________________________
___________________________________________________________.
Name: ___________________________________
An Apple a Day: Pre-Laboratory Worksheet
1. Where is DNA found? _________________________________________________________
2. What do you think the extracted DNA will look like? ________________________________
______________________________________________________________________________
3. How could DNA that is extracted from a human be used? Name 3 ways:
a. ____________________________________________________________________________
b. ____________________________________________________________________________
c. ____________________________________________________________________________
4. The human body contains about 100 trillion cells. Each of these cells contains approximately
six feet of DNA. To about how many total miles of DNA is this equivalent? Recall that 100
trillion = 1 x 1014, and 1 mile = 5,280 feet. Show your work below.
5. Why is salt used in this exercise? ________________________________________________
______________________________________________________________________________
6. Why is the Dawn dish soap (or Suave clarifying shampoo) used in this exercise?
______________________________________________________________________________
7. Why is alcohol used in this exercise? _____________________________________________
______________________________________________________________________________
8. What organism will provide DNA for extraction in this exercise? ______________________
9. A coffee filter will be used in this exercise. What do you think it will be used for? __________
______________________________________________________________________________
10. Circle the item(s) in this list that contain DNA:
Zebra fish
Shale
Ruby
Gum wrapper
Pine tree
Blood
Mosquito
Wood
Yogurt
Carrot
Orange
Cat
Quartz
Candle
Compact disc
Name: ___________________________________
An Apple a Day: Post-Laboratory Worksheet
Results, Analysis, and Conclusions
1. Describe the appearance of the suspension in the test tube. ____________________________
______________________________________________________________________________
Using the diagram below, sketch what the suspension in your test tube looked like after you
completed the last step of the exercise. Be sure to label any layers of liquid there may have been,
as well as where the DNA was:
2. Why was the salt used in the beginning step? _______________________________________
______________________________________________________________________________
______________________________________________________________________________
3. When you do dishes at home, what happens when you put dish soap in a bowl or pan with a
lot of grease (fat) in it? ___________________________________________________________
______________________________________________________________________________
4. Why was the detergent added to the banana mixture? _________________________________
______________________________________________________________________________
5. Why do you think it might be important for scientists to be able to remove the DNA from an
organism? _____________________________________________________________________
______________________________________________________________________________
6. Is there DNA in your food? ___________ How do you know? _________________________
______________________________________________________________________________
7. What do we mean when we say the DNA has been “extracted” from the banana cells? ______
______________________________________________________________________________
______________________________________________________________________________
8. List the steps of the DNA extraction process as you followed them in the exercise. What did
the banana solution look like at each step?
STEPS
OBSERVATIONS
9. How do you think the results of the exercise would change, if you had used a different
organism from which to extract the DNA? ___________________________________________
______________________________________________________________________________
______________________________________________________________________________
10. What is the purpose for extracting DNA? How might extracted DNA be used? ___________
______________________________________________________________________________
______________________________________________________________________________
______________________________________________________________________________
11. Where is DNA found? ________________________________________________________
______________________________________________________________________________
12. Why is DNA essential to all living things? ________________________________________
______________________________________________________________________________
13. Briefly explain the purpose of each of the following materials, as used in the exercise:
Plastic cup
Blender
Plastic spoon
Coffee filter
Distilled water
Detergent or shampoo
Bananas
Salt
Plastic transfer pipette
95% ethanol
Small tube with cap
Ice and ice bucket
Name: ___________________________________
An Apple a Day: Laboratory Exercise
In this exercise, you will extract DNA from bananas that have been blended with water. You will
then treat a portion of the banana mixture with detergent, salt, and cold alcohol to precipitate the
DNA from the banana solution.
Materials Needed
Blender
Two 5-ounce plastic cups
Plastic spoon
Graduated cylinder*
#2 cone coffee filters
Distilled or tap water
Dawn dish detergent**
2 peeled bananas
Teaspoon
2 pinches of table salt (iodized or noniodized)
Plastic transfer pipette or medicine dropper
Two test tubes, with caps
Test tube stand
95% ethyl alcohol***
Ice and ice bucket
Glass rod****
Post-laboratory worksheet
*Or liquid measuring cup and dry measuring spoons
** Or Suave clarifying shampoo
*** Or 91% isopropyl alcohol
****Or wooden stick, or glass Pasteur pipette modified into a hook
Procedure
Carefully read and follow the directions. Place a checkmark next to each step as you complete it.
1. Obtain an ice bucket and fill it with ice.
2. Place about 5 mL ethanol (or isopropyl alcohol) into one test tube, cap the test tube,
and place it (upright) into the ice bucket. You’ll need to use ice-cold alcohol to get
the best results for this exercise.
3. Add two peeled bananas and one cup (about 250 mL) distilled water to a blender.
Blend for 15-20 seconds, until the solution is a mixture. (The instructor may have
already performed this step, so check first.)
4. In one of the 5 oz. plastic cups, make a solution of one teaspoon of Dawn
dishwashing detergent and two pinches of table salt. Add 4 teaspoons (about 20 mL)
of distilled water or until the cup is about one-third full. Dissolve the salt and
shampoo by stirring slowly with the plastic spoon to avoid foaming.
5. Add three heaping teaspoons (about 15-20 mL) of the banana mixture from step three
to the solution -from step 4. Slowly stir the solution with the plastic spoon for 5-10
minutes. (The detergent dissolves the lipids that hold the cell membranes together,
thus releasing the DNA into solution. The detergent causes lipids and proteins to
precipitate out of the solution, leaving the DNA. The salt enables the DNA strands to
come together.)
6. Place a #2 coffee filter inside of the remaining 5 oz. plastic cup. Fold the coffee
filter’s edge around the cup so that the filter does not touch the bottom of the cup.
7. Pour the mixture from step 5 into the filter. Let the solution drain for several minutes
until there is approximately 5 ml (covers the bottom of the cup) of the filtrate, from
which the DNA will be extracted.
8. Fill the plastic pipette with solution from step 7 and transfer it to the remaining test
tube.
9. Remove the test tube of cold alcohol from the ice bucket. For best results, the alcohol
should be as cold as possible.
10. Tilt the test tube containing the filtrate to one side. Slowly pour the alcohol into the
filtrate down the side, so it forms a layer on top of the banana filtrate. Pour until you
have about the same amount of alcohol in the tube as filtrate. Alcohol is less dense
than water, so it will float on top of the banana solution.
11. Let the column sit for 2-3 minutes without disturbing it. It is important not to shake
the tube during this time, because the strands of DNA are fragile at this point. DNA is
not soluble in alcohol. When alcohol is added to the mixture, the components of the
mixture, except for DNA, stay in solution while the DNA precipitates out into the
alcohol layer.
12. You can watch the white DNA precipitate out of solution into the alcohol layer. DNA
has the appearance of white, stringy mucus. The protein and grease parts from the
banana cells stay in the bottom, watery layer of banana filtrate, while the DNA
precipitates into the top layer of alcohol. Look for DNA at the point where the alcohol
layer touches the filtrate layer, since this is where the DNA will precipitate first.
13. When good results are obtained, there will be enough DNA to spool on to a glass rod.
Or, you can retrieve some of the DNA by using a wood stick or a Pasteur pipette that
has been heated at the tip to form a hook. Once the DNA has risen into the alcohol
layer from the banana layer, the DNA will be a long, stringy molecule that clumps
together.
14. Congratulations! You have just extracted DNA! Now, answer the questions on your
post-laboratory worksheet. Clean up your workspace according to your instructor’s
directions.
Name: ___________________________________
An Apple a Day: Laboratory Exercise, Part 2
Make a Smoothie
With permission from your instructor, after you finish your extraction activity you may follow
the recipe below to make a banana-strawberry smoothie:
1. To the remaining banana mixture in the blender, add the following ingredients and blend
until smooth.
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•
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An additional peeled banana, cut into pieces
One 10-ounce package of frozen strawberries, partially thawed
One 12.3-ounce package of tofu (either soft or firm, cut into pieces)
One cup orange juice (add more for a thinner consistency)
Two tablespoons of honey (or more, to taste)
2. Share the smoothie with your classmates in 5 oz. cups. This recipe makes enough for 10
5 oz. cups.
3. Evaluate your smoothie. Why do you think the tofu is in the smoothie?