INSTRUCTIONS FOR USE OF LIQUID CULTURE VIALS
- Mycomate® Liquid Culture Vial
- Sterile alcohol swabs
- Sterile syringe and 18 gauge needle- Liquid Culture Vial
2) You'll need:
- Gas flame (a "torch type" lighter works well)
- The spores you want to inject into the liquid culture vial.
- Clean hands and a clean area to work
Injection and colonization of liquid culture:
1) If the liquid culture medium is refrigerated, remove it from the refrigerator and store it at room temperature 24 hours before use.
2) Wash your hands very well (use of antibacterial soap is optional). Dry with a clean towel (eg paper towels).
3) Remove the vial from the bag and place on a clean table in a clean area, free of debris.
4) Apply alcohol to the injection port of the Liquid Culture Vial and any other site intended for injection (eg spore vial or Liquid Culture Vial). Use a clean swab for each product to be moistened, in order to reduce mutual contamination. Do not exhale directly over injection sites.
5) Remove the syringe and needle from the bag. Remove the plastic cover that protects the needle (turn clockwise and pull).
Do not allow the needle to come into contact with any object, otherwise it will need to be re-sterilized. If a syringe from any other supplier is used, it must also be sterilized.
Needles can be sterilized with a flame. You want the needle to be red hot, being careful not to overheat and melt the area where the needle is attached to the plastic. Always let the needle cool before injecting it. (See photo a).
6)If a colonized liquid culture or spore vial is used, shake vigorously to mix the suspension. Hold the vial at an angle, inject with the syringe, and withdraw the desired amount of suspension. (See photo b).
7)Inject several drops of culture suspension (ie, colonized fluid from the culture vial) or between several drops and 1 mL of spore suspension (depending on the density of the spore solution) into the Liquid Culture Vial. Withdraw the needle very slowly, otherwise the pressure may cause some of the liquid to spray out—which is not a problem. (See photo c).
8) Clean the injection port of the Liquid Culture Vial with a clean swab moistened with alcohol; let dry, and store upright in a zip-lock plastic bag.
9) Vigorously shake the vial once a day for ten seconds and store away from sunlight. Incubate fungal species at the appropriate temperature (generally 23º to 27º C). A lower temperature delays the germination and colonization time of the spores. However, a temperature that is too high (above 27º C) increases the risk of contamination. (See photo d).
Note: Temperature requirements depend on the species.
10) Growth is similar both with spores and with Liquid Culture. However, growth with spores takes longer than with the latter.
Spore suspension: Spores generally take 24 to 72 hours to germinate, and colonization of the liquid medium should be observable within 3 to 7 days growing in the lower two-thirds of the vial.
Note: MycoMate® Liquid Culture Vials require very few spores for growth.
Liquid Culture: Growth is evident within 24 hours of transferring mycelium from a colonized Liquid Culture vial to fresh Liquid Culture vials. Fresh vials are generally colonized after several days (depending on species, variety, and temperature).
Observation is most effective when the vial is carefully held up to the light, avoiding shaking. The sediment in the culture medium can be agitated if you are not careful enough, making it more difficult to identify the growth (this is not a problem, since after a while the sediment will settle back to the bottom). White strands of mycelium (fungus) will float to the surface as you lift the vial. There should be a little cloudiness around the mycelium, but the entire vial should not be cloudy. The area around the cloudiness of the mycelium should be reasonably transparent. If the vial is lifted slowly and yet appears completely opaque, it may be contaminated. Let the vial sit undisturbed for several hours and check again to see if it is sediment from the bottom that has obscured the vial. This can be used as soon as there is healthy growth or after one to two weeks when the vial shows increased colonization.
Note: When between one and two thirds of the vial has been colonized it should be used or stored under refrigeration for later use (see below).
11) When colonization is complete the vial can be used to inoculate other MycoMate® Liquid Culture Vials (repeating steps 1-10), MycoMate® Kits for Edible Fungi or Sclerotia (repeating steps 1-6)
MycoMate® Liquid Culture Vials can be used for the storage of both short-term and long-term cultures, including multispore, isolated or cloned varieties. One Liquid Culture Vial can be used to produce thousands more. Once the vial has been inoculated, and the mycelium colonizes between 30% and 65% of the liquid medium, it can be stored under refrigeration (between 2º and 8º C) for several months (or more) without perceiving effects. harmful. Store upright in two or three hermetically sealed plastic bags. Shaking of the cultures is not required during this period.
Save valuable time: When liquid culture is required, simply remove the vial from the refrigerator and use.