The Best Agar Recipe for Mycology (Complete Tek)

Exact Recipe (500 mL ≈ 18–22 plates)

• 10 g agar-agar powder
• 7.5 g light malt extract (LME)
• 0.25 g soy peptone
• 0.25 g powdered nutritional yeast (optional)
• 500 mL distilled or RO-filtered water
• Food coloring, 1–3 drops (optional; aids visibility)

Why this formula? It’s nutrient-balanced for fast, robust growth while staying visually clean, so you can easily see mycelium and spot contaminants.

Supplies & Gear

Containers & Tools

• 500 mL glass media bottle or heat-safe Mason jar (with screw cap)
• Aluminum foil (to cap/cover during sterilization)
• Pressure cooker capable of 15 PSI (Presto/All American)
• Trivet/rack for inside the pressure cooker
• Infrared or probe thermometer (for pouring temperature)
• Still Air Box (SAB) or laminar flow hood
• Disposable 90 mm sterile Petri dishes (or reusable glass plates)
• Gloves, face mask, 70% isopropyl alcohol, paper towels
• Small funnel (optional, for neat pouring)
• Parafilm M or micropore tape for sealing plates (optional but recommended)

How Much This Makes (Fill Volumes & Plate Counts)

Standard fill is 20–25 mL per 90 mm plate (≈ 1/8″ depth). Expect minor losses from steam & bottle residue.

Step 1 — Mix the Medium

1. Weigh all powders into a dry media bottle or Mason jar.
2. Add 500 mL water. Swirl vigorously until no dry clumps remain (a few fine bubbles are okay).
3. Add 1–3 drops food coloring if desired. Swirl. (Adding color now mixes evenly and avoids extra handling later.)
4. Wipe the threads, cap loosely, then wrap the cap with foil (or just foil over a Mason jar) to protect from drips.

Step 2 — Sterilize

1. Place a trivet/rack in the pressure cooker and add water per manufacturer (usually 1–2 inches).
2. Set the bottle on the rack (never on bare metal). Leave the cap slightly loose so steam can escape.
3. Vent a steady jet of steam for ~10 minutes, then bring to 15 PSI.
4. Hold for 30–40 minutes, then turn off heat.

Important: Depressurize in Clean Air

If you have a flow hood, move the pressure cooker in front of it before pressure drops to zero and let it cool/depressurize there. Opening the PC under clean airflow reduces the chance of unfiltered air being pulled into the bottle as it cools (negative pressure effect).

Step 3 — Cool to Pouring Temperature

• Target pour range: 115–125°F (46–52°C). Below ~110°F it may start gelling in the bottle; above ~130°F promotes condensation in plates.
• Check temperature with a thermometer. If you don’t have one, wait ~25–35 minutes after the jiggle stops, then test a tiny dribble—should be hot but not steaming vigorously.
• Keep the cap just snug while cooling (not fully tight) to avoid vacuum lock.

Step 4 — Pour Plates (Stack Method)

Set Up Your Sterile Area

• Work in a SAB or in front of a flow hood. Spray/wipe surfaces with 70% IPA. Wear gloves and a mask.
• Arrange stacks of 5–10 plates. Label the bottoms (agar side) now (date, formula, color).
• Warm, dry room helps; avoid fans/vents.

Pouring Technique

1. Bring the bottle into the sterile field. Tighten the cap fully, remove foil, then crack cap slowly to equalize pressure.
2. Work top-to-bottom within each stack: slide the top lid just enough to pour; don’t fully remove it.
3. Pour 20–25 mL into each plate (≈ one thin layer covering the base). Close immediately.
4. Repeat down the stack. Keep movements slow and deliberate to avoid stirring air.

Minimizing Condensation

• Pour at 115–125°F. Too hot = more steam & condensation.
• Pre-warm plates 10–15 minutes in the sterile area (or on a warm, clean surface) so plastic isn’t cold.
• After pouring each stack, rest lids slightly ajar for 15–30 seconds to vent steam, then close fully.
• Let plates set in tall stacks (retain gentle warmth) on a flat, vibration-free surface.
• Once solid (~10–20 min), flip to inverted storage (agar up, lid down) to keep condensation off the surface.

Step 5 — Curing, Storage & Shelf Life

• Let poured plates “cure” closed for 12–24 hours so micro-condensation dissipates.
• Seal edges with Parafilm (one wrap around the rim) or micropore tape.
• Room temp: clean, sealed plates last 2–3 months.
• Refrigerated (not required): up to 4–6 months. Bag to avoid fridge humidity; let warm to room temp before use to prevent surface wetness.
• Always inspect: if you see foggy growth, pigmentation, or droplets/puddles on the agar, discard.

Using Your Plates

• Work sterile (SAB/flow). Flame-sterilize scalpel; cool on sterile agar edge or a cool agar wedge before touching culture.
• Label transfers clearly by date and passage (T1, T2…).
• Incubate most gourmet species at 70–75°F (21–24°C) until growth is visible.

Alternatives if You’re Missing Supplies

• No peptone? Omit it. Plates still work; growth may be slightly slower.
• No nutritional yeast? Skip it (it’s optional). Brewer’s yeast powder is a functional substitute.
• No LME? Use potato flakes (make potato infusion) or dextrose-based PDA. Expect darker color with PDA.
• No media bottle? Use a Mason jar with a metal lid; keep it slightly loose and foil-covered in the PC.
• No thermometer? Let the bottle sit ~25–35 minutes after pressure drops to zero; begin pouring when it’s hot but not steaming vigorously.

Pro Tips & Common Pitfalls

• Plates too firm? Reduce agar to 9 g per 500 mL next batch.
• Plates too soft? Increase agar to 11–12 g per 500 mL.
• Cloudy plates right after PC? Fine particles settle as it cools; clarity improves. Persistent murkiness may be over-cooked sugars—shorten PC time to 30–35 minutes.
• Droplets on lids? Store inverted. If surface gets wet, leave plates sealed at room temp for 24–48 h to dry before use.
• Bottle gelling before you finish? Gently re-melt by setting the bottle (cap loosened) in a hot-water bath (~60–70°C) and continue pouring.