Analyzing the Products of DNA:Chromatography of Fruit Fly Eye Color

Name: ____________________

Eye pigmentation in fruit flies (Drosophila) is genetically controlled. Drosophila have genes that encode pigment-forming enzymes that make the following pigments from pigment precursor molecules.

How fruit fly eye color is made:

Some questions to get you thinking about today's lab:

What is a mutant? What is a mutation?

How can we tell when an organism is mutant or has a mutation?

Materials

• Whatman filter paper #1 (others will work as well), cut in 4 inch squares
• 1:1 solution of isopropanol (2-propanol) and ammonium hydroxide
• glass or metal rods (1 per group)
• Drosophila, wild type and mutants
• pencils (1 per group)
• glass beakers (1 per group)
• aluminum foil
• UV lamps (long wavelength)
• paper towels

Procedure

1. Draw a pencil line across one end of the filter paper 1 centimeter from an edge
2. Fold ends of paper in so it can stand independently with the line across the bottom.
3. The next step is to mush 1-2 flies on the pencil line, so each member of the group should mark where on the line they will mush their flies. Which flies do you want to use?
4. Once everyone has written which fly they are putting where, mush 1-2 flies on each spot. Why can we mush the whole fly and not just the eyes?
5. Stand paper with pencil end down in a beaker with just enough isopropanol/ammonium hydroxide solution so that the pencil line is NOT sitting in the solution. What does the isopropanol/ ammonium hydroxide solution do? Why don't we want the pencil line to be soaking in the solution?
6. Cover the beaker with aluminum foil. Why?
7. Let the chromatograph run until the solution reaches the top, soaking the paper entirely (usually takes about 2 hours, can run overnight). What would happen if we took the paper out of solution before then?
8. Take the paper out, letting it dry on a paper towel covered in aluminum foil (usually takes about 1 hour).
9. Examine your Drosophila eye pigments!

Examining the Drosophila Eye Pigment Data

Use a UV light to examine your filter paper chromatogram (be sure to wear protective eye wear if you are using short wavelength UV light!).

What does the pigment look like? Draw a picture of your chromatogram.

Mark in the table which mutants have or are missing which eye pigments.

Pigment Table

Pteridine (color)	Wild Type	Mutant 1	Mutant 2	Mutant 3	Mutant 4
Isosepiapterin (yellow)
Biopterin (blue)
2-amino-4-hydroxypterin (blue)
Sepiapterin (yellow)
Xanthopterin (green-blue)
Isoxanthopterin (violet-blue)
Drosopterin (orange)

What can you conclude about your Drosophila mutants from your eye pigment chromatogram data?

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BIOTECH Project Department of Molecular and Cellular Biology The University of Arizona September 15, 2000 Designed by: Erin Dolan Nadja Anderson, Ph.D. nadja@email.arizona.edu http://biotech.biology.arizona.edu