Oct 17, 2023
Soil extract for cocultures host-parasites media
Forked from Soil extract for algal media
- Roscoff Culture Collection1,
- Estelle Bigeard2,
- Laure Guillou2
- 1Station Biologique, CNRS-Sorbonne Université, Roscoff France;
- 2Station Biologique de Roscoff, France
Protocol Citation: Roscoff Culture Collection, Estelle Bigeard, Laure Guillou 2023. Soil extract for cocultures host-parasites media. protocols.io https://dx.doi.org/10.17504/protocols.io.rm7vzxkq2gx1/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 18, 2023
Last Modified: October 17, 2023
Protocol Integer ID: 87926
Funders Acknowledgement:
ANR AQUARADOX, PARALEX, EPHEMER
Grant ID: SU & CNRS
Abstract
How to prepare soil extract
Materials
Equipments
Autoclave
Laminar flow cabinet (or biosafety cabinet)
Vacuum pump
Magnetic hot plate stirrers
Materials
Glass fibre prefilters (Millipore, AP1507500)
Filtration unit Stericup GP Merck Millipore, pore size 0.22 µm (Millipore, S2GPU02RE)
Before start
Please refer to our general recommendations to grow cultures :
https://www.protocols.io/private/A48906DC1374AD6281495CB86A8F092F
Soil sampling
Soil sampling
Soil is sampled in a pine wood, located in Brittany, France.
To find a correct sampling area in your country, try to find a location devoid of pesticides and pollutants. It should be also a place where you know that you can collect the soil again and later in order to always use the same soil, especially if this one worked.
Collect the soil in plastic boxes.
Back to the laboratory, plastic boxes are placed at 37°C for 12 to 24 hours to let the soil dry. The soil must be dry before use, otherwise the boiling step should be longer.
Preparation of the soil extract solution (25g/l)
Preparation of the soil extract solution (25g/l)
Weigh 30g of dried soil and place it in a 4L glass erlenmeyer.
Add 1200ml of MilliQ water to the soil.
Close the erlenmeyer with a aluminium foil.
Heat at 100°C with pemanent mixing during 1 hour to obtain an efficient boiling.
Adjust this time depending on the volume (more volume, more time).
Let cool.
Filtrate on 0.2 µm, using a glass prefilter and a pump vacuum.
As the prefilter will be rapidly clogged, better to add the boiled soil 100mL per 100mL.
If this happens, replace the glass prefilter, then the filter.
Transfer the solution in one or several glass bottles.
Autoclave at 121°C for 20 minutes.
Place the bottle of the filtered-boiled soil under a safety cabinet, and let it cool.
The day after, when the medium is at room temperature, place it at 37°C for 24 to 48 hours.
Then autoclave again at 121°C for 20 minutes.
This step allows to reactivate spores (fungi etc) contained in soil which have not been destroyed by the first sterilization, and to kill them during the second one.
Add 50ml of soil extract per 1 L of medium.