Neuroscience
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Taste Preference Assay for Adult Drosophila
Chapters
Summary September 8th, 2016
Taste is an important sensory process which facilitates attraction to beneficial substances and avoidance of toxic substances. This protocol describes a simple ingestion assay for determining Drosophila gustatory preference for a given chemical compound.
Transcript
The overall goal of this procedure is to determine the relative preference of adult Drosophila from one chemical tastant over another. This method can help answer key questions in the Drosophila taste field. Such as which genes are necessary for selecting between sweet compounds or potentially toxic, bitter compounds.
The main advantage of this technique, is that it is straightforward and does not require any specialized instrumentation. Demonstrating the procedure will be Andrew Bantel, a technician from my laboratory. To begin the experiment, saturate a cotton ball with water at the bottom of a standard fly vial.
Next, collect flies in sets of 100 animals on a CO2 pad and then add the flies to a prepared vial. Use a cotton ball to secure the vials closed. Then, place the vials on their side in an environmentally controlled incubator.
Prepare the controlled tastant of one millimolar sucrose, by combining 10 microliters of 100 millimolar sucrose solution, 13 microliters of red food coloring, and 977 microliters of water. Next, prepare the experimental tastant of five millimolar sucrose. Then, create the assay chambers, by obtaining a 100 millimeter by 15 millimeter plastic petri dish, and placing three microliter drops of controlled tastant as close to the edge of the plate as possible at the 12 o'clock position.
At the three o'clock position, place three 10 microliter drops of experimental tastant as close to the edge of the plate as possible. Place another three drops of the controlled tastant at the six o'clock position. Then, place another three drops of the experimental tastant at the nine o'clock position.
Next, empty one vial of 100 starved flies onto a CO2 pad, just long enough to anaesthetize all animals. Brush the animals into the middle of a prepared assay chamber, and cover it with the dish lid. Place the assay chamber into an opaque cardboard box.
Then, label the outside of the box with the condition and genotype being tested. Move the entire setup into an incubator. After two hours, place the assay chambers still contained within cardboard boxes, directly into a negative 20 degree Celsius freezer until they are ready for quantitation.
Allow a single assay chamber to warm up to room temperature. Under a dissection microscope, use a brush to group the animals based on the color of their abdomen. Red, blue, or purple.
Record the number of animals in each grouping. This graph shows the effects of varying concentrations of blue dye, while maintaining a constant concentration of red dye, revealing an optimum combination of 1.3%red, to 1.0%blue. Drosophila are naturally attracted to high sucrose concentrations, as seen by a preference index value near one, when given the choice between five millimolar sucrose and one millimolar sucrose.
This preference was reversed with acid, which dropped the preference index to near zero. While performing this procedure, it's important to remember to test the food coloring additives, were there affects on the preference assay? The concentrations of each dye should have no impact on the performance index.
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