# Fruit flies

This is a brief course of how to grow fruit flies with limited resources. The methods presented are only one example. In other words, this it not a source for a university research but instead aims for a level of natural-science in high school. The course haven’t either included what sorts of experiments are interesting to do, instead, puts focus on how to grow, manage and take care of them.

## Fruit Fly – Drosophila Melanogaster

A fruit fly lives for approximately 14 days. During this period it goes from egg, larvae, cocoon till a fly. It takes around ten days for the fly to develop from the day when the egg is laid. Notable, females are ready for courting around 10 hours after they have hatched.

### Different flies

* The female fly is larger than the male and have a greyish rear part.
* The male fly is smaller than the female and their rear part is black. As well, the male have a constellation of spiky hair around the genitalia used to attached the female during the mating process.
* You can perceive a female virgin by its whiter rear part.

## Short basics of cultivation

* Try to have around 20 flies in a vial.
* Always mark a new vial with a unique number and the date of transfer.
* Transfer flies every 2-3 day.
* Food never has to be replaced, if a old vial runs out of food, replace the whole vial.
* Keep track on how you transfer your flies from old vials to new vials. Best way to do it is to continuously update a schedule.
* Keep in mind that you will have to spend more time than intended in the beginning, and over all, growing fruit flies may be time-consuming.

## Environment

It’s important to keep the environment stable. The easiest way is to grow the flies in a normally tempered room. Larger rooms will give a more sustainable environment. Keep in mind that temperature and humidity can vary largely in environments you don't expect it to (night time, day time, sunny or cloudy), therefore, investigate the room before starting to grow the flies.
It could be a good idea to place a glass of wine close to the cultivation, since some flies will escape. The flies like the smell of the wine and drown when they land in the liquid.
* The optimal temperature to grow flies in is 25°C - 28°C. Flies may live at 20°C or under, but the flies reproduce faster if it is warmer. On the other hand, temperatures above 29° - 30° will make the male flies sterile
* Humidity should be around 50% - 60% outside the vials, if air easily can pass through the seal. There shouldn’t be any liquid water on the wall of the vial, because of the risk of “drowning” the flies. If the water from the food (see below) is not enough to sustain the humidity level, small bottles of water can be placed close by the vials (doesn't work if growing the flies in a room).

## Case, vials and seals

Depending on your intentions on experiments, or if there is no well tempered room to place the flies, you may have choose to place the flies in a bigger case. The more volume the easier it is to keep the humidity and the temperature at a desired level.
* The case shouldn't be much smaller than 30*20*20 centimeters. Moreover, the case preferably should be transparent and allow some air exchange with the outside.
* The best and easiest vials to use are the one that can be bought.
* To the standard vial you can by foam rubber seals which are the easiest to use. Otherwise, pressing together a piece of cotton is a good solution.

## Sterilize materials

Depending on the level of research, you should sterilize the materials and most importantly the vials.
* Prepare a container filled with distilled water, something to heat the water with (a burner and a tripod from the chemistry lab), vials and tools (e.g. a suitable pairs of forceps). Hand wash the container, vials and the tools before use.
* Heat the water to a boiling temperature.
* Clean the tools. A recommendation is to have two containers, one smaller for cleaning the tools and enough, in order to save the time it take to get the water boiling.
* Heat new distilled water for the vials.
* When boiling, put in as many vials as there is room for 5 seconds.
* Place to dry.
* Put away the vials in a closed container. They can be stored for a longer time.
Seals should not be sterilized, since they doesn't endure the heat of boiling water.

## Food

The food is vital for the flies and influences the environment and the relative time of reproduction. The food is the most important medium which is to keep the humidity at a desirable level. Therefore, the amount of liquid in the food and its consistency are important to have in mind. Do evaluate how the food is working for your usage and try to optimize. There are many different recipes that are to be found on the internet, below is an example of a bought medium mixed with water.
* Prepare vials, a big container, the food medium, jest, viscous syrup, some small spoons and something to stir with.
* Work by a water tap and hand wash the tools.
* Fill the container with your choice of food medium and a spoon of jest per five decilitre food medium. Mix.
* Pour in the right amount of water, but replace one tenth of the water with syrup to get a firmer consistence. Stir.
* The consistence of the food should be solid, but if you gently press on it with a finger you should be going down/in to the medium, and get wet.
* Portion the food into the vials, about 1.5 centimeters of the vials height from the bottom should consist of food. Press the down the food into one solid mass.
Things to keep in mind
* You save time if you make much food at the same time, but if you do not use the food filled vials directly, they should be stored in a refrigerator.
* If the food tends to fall down during transfer (see below), you could add more syrup to make it stickier. Though, don't replace the water entirely, since the food in that case will become too dry.

## Transfer of flies from vials to vials

Transfer flies to a new vial every 2-3 day. Mark the new vial and prepare it with food before transfer. Have a new seal ready. This process is easier done in pairs.
* Bump the vial with the flies on to the table gently, enough to keep the flies in the bottom of the vial. Try to tilt the vial and bump the edge of the bottom on to the table, so the flies do not fall directly into the food, where they may get stuck.
* Remove the seal so the opening is all clear.
* Quickly put the new vial on top of the old one, leaving no gap for escape.
* Turn the two vials upside down, meanwhile keeping the two vials together.
* Bump the vials to the table so the flies fall down in to the new vial.
* Part the vials and (together with your friend) quickly seal the vials. Be aware of flies that still may be in the old vial.
Things to keep in mind
* Food may fall down during transfer when bumping the vial, a devastating catastrophe. Therefore, try do make the process/bumping as quick and gently as possible.
* If using cotton seal: If some of the cotton from the seal sticks to the vial’s wall, creating a “net/beard”, remove it quickly before turning the vial upside down, meanwhile bumping the vial into the table in order to keep the flies in place.
* When sealing, be aware of not crushing the flies with seal
* If you are doing any experiments where it is important to keep track of where the flies are transferred, you should organize a schedule before starting to grow the flies.

## Exterminate flies and reuse the vials

The food in older vials may after a while look really nasty (grey and very wet). Preferable is to kill the flies and reuse the vials. There is numerous ways to kill flies. It is important to try to keep the population under a decided amount, otherwise the flies tend to grow into uncontrollable numbers. The best way to kill the flies, and be sure there is no living larvae left in the vial, is to simply but the vial into the freezer. Afterwards, clean and sterilise the vials thoroughly.

## Put flies to sleep (with CO2)

When you need to count the flies accurately, transfer them in a certain number or divide by any attribute (e.g. male or female). WHOP The procedure following is designed by a shortage of founds, there are much better methods to use. You definitely have to be more than two for this to work.
* Prepare the CO2 tank, plates with high edges (preferable transparent material) , soft brushes to move around the flies and if needed new vials with seal.
* Gently push the slang from the CO2 tube into the seal to pour the gas into the vial. Be careful so the seal doesn’t slide in and crush the flies.
* When flies are sound asleep, transfer them to the plate.
* The flies will eventually wake up, therefore pour in CO2 onto the plate when needed. Since the CO2 is heavier than air it will stay in the plate for a while.
* Do what you have to do with your pencils, but work fast.
Things to keep in mind
* Flies sometimes wake up faster then predicted, always be ready with the CO2.
* Try to give the flies small but many doses of CO2, since too much at a time will suffocate them.
* Some flies will inevitable escape, therefore, be in another room than the cultivation.
* Do not try to enclose the flies into any plate or container, since, they eventually will wake up and it will be very difficult to put them back to sleep or in a veil.

## Literature and sources

* http://biology.arizona.edu/sciconn/lessons2/Geiger/intro2.htm#disease
A schoolteacher presenting his methods and choice of project/experiment.
* http://flybase.org/
A database of the Drosophila genes and genomes.
* http://en.wikipedia.org/wiki/Drosophila_melanogaster
An over-all article.
* http://www.is.wayne.edu/mnissani/PAGEPUB/FLIES.HTM
Course material and setups.

### Source of images

* Figure I: Kevin Edwards, 2012-03-11, http://bio.illinoisstate.edu/kaedwar/other/Sexing_Drosophila.shtml
* Figures II – V: illustrated by Henrik Almquist

## Acknowledge

This document is based on information acquired through experience and sources found under the title Literature and sources. Additionally, there has been a contact with a section at Stockholm University's which work with fruit flies. This is the product produced after a project work at the high school of Rudbeck in Sweden during the autumn 2011 and spring 2012. The team consisted of three members: Gustav Nordqvist, Jonathan Nilson and Henrik Almquist.
Do not hesitate to contact us if there are any questions or remarks. Email: me@gmail.com