This is a protocol preprint. Preprints are preliminary reports that have not undergone peer review. They should not be considered conclusive, used to inform clinical practice, or referenced by the media as validated information.
Method Article
Preparation of a holidic medium for Drosophila melanogaster
Matt Piper
Piper Lab
Corresponding Author
License:
This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full License
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.
Keywords
drosophila, holidic diet, nutrition
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:
- purchase iungredients according to order sheet (Table 1)
prepare each of the stock solutions using the qauntities and according to the directions shown in Table 2, Table 3 and Table 4
MEDIA
Preparation of the medium is performed in two stages.
(also see Table 5: example working recipes)
I. before autoclaving
- in a 1l, autoclave-proof, glass bottle, mark on the side the level of the target pre-autoclave volume (=1 litre minus the sum of the ingredients to be added after autoclaving)
- add magnetic stirrer flea to bottle
- to milliQ water, add sucrose and low solubility amino acids (L, I, Y)
- add stock solutions for buffer base and metal ions
- stir well (may not fully dissolve at this stage)
- add cholesterol stock solution (solution will turn cloudy)
add agar
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)
- allow solution to cool at room temperature with stirring to ~65 °C
- add sterile stock solutions in order: amino acids, nucleosides/choline/inositol mix, vitamins and preservatives
- using sterile tubing, dispense the solution into sterile vials
- cover the vials and allow to cool for 90 min at room temperature
- stored at 4 °C until use
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
This is a list of supplementary files associated with this preprint. Click to download.
- Stocksolutionsbeforeautoclaving.pdf
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.
- Stocksolutionsafterautoclaving.pdf
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.
- Stocksolutionsafterautoclavingaminoacids.pdf
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.
- exampleworkingrecipes.pdf
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.
- reorderlist.pdf
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.
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Version History
CURRENT STATUS: Posted
Version 1
Posted 18 Nov, 2013
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Subject Areas
Biological techniques
Associated Publications
A holidic medium for Drosophila melanogaster
by Matthew D W Piper, Eric Blanc, Ricardo Leitão-Gonçalves, +11
Nature Methods (18 November, 2013)
Method Article
Preparation of a holidic medium for Drosophila melanogaster
Matt Piper
Piper Lab
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 18 Nov, 2013
No community comments so far
Metrics
Comments: 2
HTML Views: ...
Subject Areas
Biological techniques
License:
This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full License
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:
- purchase iungredients according to order sheet (Table 1)
prepare each of the stock solutions using the qauntities and according to the directions shown in Table 2, Table 3 and Table 4
MEDIA
Preparation of the medium is performed in two stages.
(also see Table 5: example working recipes)
I. before autoclaving
- in a 1l, autoclave-proof, glass bottle, mark on the side the level of the target pre-autoclave volume (=1 litre minus the sum of the ingredients to be added after autoclaving)
- add magnetic stirrer flea to bottle
- to milliQ water, add sucrose and low solubility amino acids (L, I, Y)
- add stock solutions for buffer base and metal ions
- stir well (may not fully dissolve at this stage)
- add cholesterol stock solution (solution will turn cloudy)
add agar
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)
- allow solution to cool at room temperature with stirring to ~65 °C
- add sterile stock solutions in order: amino acids, nucleosides/choline/inositol mix, vitamins and preservatives
- using sterile tubing, dispense the solution into sterile vials
- cover the vials and allow to cool for 90 min at room temperature
- stored at 4 °C until use
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
2 comments flagged for moderation
If you posted a comment and think it was mistakenly removed, please contact customer support at [email protected].
Associated Publications
A holidic medium for Drosophila melanogaster
by Matthew D W Piper, Eric Blanc, Ricardo Leitão-Gonçalves, +11
Nature Methods (18 November, 2013)