Sep 27, 2023
Drosophila Locomotion Protocol
- Mel Feany1,2
- 1Harvard Medical School;
- 2Brigham and Women's Hospital
Protocol Citation: Mel Feany 2023. Drosophila Locomotion Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.4r3l226p4l1y/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: September 27, 2023
Last Modified: May 31, 2024
Protocol Integer ID: 88491
Keywords: ASAPCRN, Drosophila, Locomotion, Screening
Funders Acknowledgement:
MJFF
Grant ID: 000301
Abstract
This protocol is used to assay the locomotion of flies as a screening tool.
Day -10: Set up crosses
Day -10: Set up crosses
Set up fly crosses as needed.
Day 0-6: Collect progeny
Day 0-6: Collect progeny
~10 days after setting up the crosses, progeny will begin to eclose. Once daily, collect those that are desired, discarding any undesired genotypes. Flies should be collected in vials with 9-14 flies per vial.
6 replicates per genotype is desirable for the assay, so once 6 vials (of 9-14 flies each) have been collected, the crosses may be discarded even if they are still producing additional flies.
Note: Flies are sensitive to the effects of anesthesia, so collecting them at the time of eclosion is the only time they should be exposed to CO2.
Day 1-10: Aging flies
Day 1-10: Aging flies
Flies should be flipped (without anesthesia) to new food every 2-3 days while aging and stored at 25 °C in an incubator.
Day 10*: Locomotion assay
Day 10*: Locomotion assay
1m
1m
Supplies needed:
Timer
Empty vials and buzz plugs
White paper
iPhone or iPod, or alternate recording device, optional
*Day 10 is a standard time point but the assay can be performed at any desired timepoint and will show age related decline in locomotion over at least the course of 3-20 days.
Perform assay in groups of 4-7 vials at a time when possible. For example, if 30 vials are to be assayed on one day, divide them into 5 groups of 6 or 6 groups of 5.
Flip flies to new polystyrene or polypropylene vials, without food, plugged with a Buzz Plug. Write the genotype or code on the bottom of the vial so that it does not interfere with seeing the flies when the tube is on its side.
Allow flies to acclimate for 00:01:00 before beginning.
1m
Start Timer. Briefly but firmly tap the first vial 3 times, strong enough to knock the flies to the bottom on the vial, then immediately lay it on its side on top of a white piece of paper and note the time on the timer. Wait 15 seconds, watching the time count down on the timer.
After the 15 second delay, record a 3 second video of the flies moving in the vial using an iPhone or iPod or equivalent.
(As an alternative, counts can be performed manually).
After the video is recorded, put the vial upright and set aside for now.
Repeat steps 8-10 for each additional vial, until each vial has been recorded 1 time. Then start back again with vial #1, completing a total of 6 sets of recordings.
At the end of the assay flies are anesthetized and fixed or frozen as needed for downstream assays.
Analyze movement data in a blinded fashion.