Technique for pouring of agar-solidified nutrient medium into Petri dishes.

Materials

  • Melted C-Fern Medium from one or more of the following sources
    • Bottled C-Fem Medium
    • Previously prepared agar-solidifed C-Fem Medium
      (Previously prepared or bottled medium must be melted well in advance. To speed melting, vigorously shake the bottle to break up the medium just prior to placing bottle in water bath. To melt medium, loosen caps on containers and place containers into a hot water bath such that the water level is only just above the level of the medium in the container. Do not submerge the container. A cover on the water bath helps the medium to melt faster. In a boiling water bath (100°C or 212°F), the melting time for medium is about 15 min for 160 mL and 45 min for 400 mL.)
    • Newly prepared and autoclaved agar-solidifed C-Fem Medium
  • Sterile Petri Dishes, 60 x 15 mm
  • Clean Area for Pouring Dishes (possibly, a sterile bench or laminar flow hood)

Methods

Medium should be poured into dishes at least 4 hr in advance by students or the instructor to allow time for it to cool to room temperature and solidify completely.

If a sterile bench or laminar flow hood is not available, medium should be poured in a clean area free from drafts and traffic. Basic C-Fem medium lacks carbohydrate sources (sugars) so contamination is rarely a problem. Prepare the area by wiping it down with 70% ethanol or isopropanol or with a damp clean sponge.

Open a sleeve of 60-mm Petri dishes by cutting through the end of the bag with scissors. Save each Petri dish sleeve for temporary storage of dishes. To pour dishes, be sure that medium is completely melted, gently swirl container to be sure that medium is thoroughly mixed, remove the cap, then tilt the lid of a petri dish upward just enough to permit pouring of medium into the dish. Fill dishes about ¾ full, i.e. about 15 mL for a 60-mm Petri dish. Do not underfill dishes; 160 mL of medium should prepare about 10 petri dishes; 400 mL about 25 dishes. Carefully replace petri dish lid and allow dishes to cool undisturbed. Do not jar dishes. Dishes may be poured in stacks of 5-10, starting with the ottom and working upward. Condensation that may form on Petri dish lids during cooling is minimized if dishes are poured and cooled in stacks. Once dishes have cooled and the medium has  solidified, they may be returned to the plastic sleeves. Seal the plastic sleeves with tape and store room temperature prior to their use.