Method Article
Preparation of a holidic medium for Drosophila melanogaster
https://doi.org/10.1038/protex.2013.082
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We describe a novel holidic medium for Drosophila melanogaster that has been optimised for adult fitness traits. Development is also possible on this medium, albeit at a reduced rate.
On this medium, adult egg laying, lifespan and activity patterns are no different from those of flies on a sugar yeast diet.
Drosophila melanogaster feeds on fermenting fruit.
Here we describe a synthetic, fully chemically defined (holidic) medium that it is sufficient for fruitfly development, albeit at a reduced rate. When adult flies are maintained on the diet, they are phenotypically similar to those kept on a natural yeast based (oligidic) diet. The holidic medium can produce more stable experimental outcomes than oligidic media, thus potentially improving inter-laboratory comparability. This medium also offers the opportunity to investigate the effects of subtle nutrient manipulations that could not be achieved otherwise.
see table of reagents and order numbers in Table 1
silicone tubing
autoclave
autoclave-resistant bottles
magnetic stirrer
sterile syringes
0.22 micron syringe-mounted filters
IN ADVANCE:
MEDIA
Preparation of the medium is performed in two stages.
(also see Table 5: example working recipes)
I. before autoclaving
II. autoclave at 120 °C for 15 min
(at the same time, autoclave dispensing tubing and any glass vials or bottles to house the medium)
III.
(all steps with constant stirring)
Stock solution preparations:
~30 - 60min each
medium preparation before autoclaving
~30 min
autoclaving (variable depending on model)
~90 min
medium mixing after autoclaving
~15 min
dispensing medium
~30 min
time to cool
90 min
MEDIUM DOESN'T SET:
Try sterilising the buffer base separately from the medium and adding it after autoclaving.
In addition to the authors, the following people contributed significantly to the development of the medium:
Eric Blanc, Ricardo Leitão-Gonçalves, Mingyao Yang, Xiaoli He, Nancy Linford, Matthew Hoddinott, Corinna Hopfen, George Soultoukis, Christine Niemeyer, Fiona Kerr, Scott Pletcher, Carlos Ribeiro
none
Table 1 Ordering list for holidic diet components A list of holidic medium components with order numbers. These are supplied in an attempt to standardise how people make the media and enhance replication of data between laboratories and trials.
Table 5 example working recipe These recipes describe the ingredient list and quantities required to make up holidic media for HUNTaa and Yaa amino acid mixtures. Note, there are three recipes for each corresponding to different final amino acid levels in the media. Modifying these levels will alter adult fitness measures.
Table 4 Stock solutions part 3 (added after autoclaving) These are amino acid stock solutions. Note, there are two tables here, each containing all 20 amino acids, but in differet ratios. The Yaa ratio is that most similar to what is found in yeast.
Table 3 Stock solutions part 2 (added after autoclaving) These tables contain the stock solutions to be added after autoclaving. In most cases they are filter sterilised using a syringe-mounted filter. They are added after autoclaving to avoid nutritional loss upon heating.
Table 2 Stock solutions part 1 (those added before autoclaving) These stock solutions are to be made in advance of medium preparation. This batch are added to the medium before autoclaving.
This protocol has been posted on Protocol Exchange, an open repository of community-contributed protocols sponsored by Nature Portfolio. These protocols are posted directly on the Protocol Exchange by authors and are made freely available to the scientific community for use and comment.
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