Agar Work
Agar recipes, transfers, strain-specific media, and techniques for isolating clean genetics.
What Is Agar and Why Use It?
Agar is a jelly-like substance derived from seaweed that, when mixed with nutrients and water, creates a solid growth medium for mushroom mycelium. Working with agar is one of the most important skills in mushroom cultivation because it gives you control over your genetics and lets you verify culture cleanliness before committing to grain.
Key Reasons to Use Agar
| Purpose | Description |
|---|---|
| Isolating genetics | Multispore syringes contain thousands of genetically unique spores. Agar lets you isolate a single strong strain through successive transfers, giving you consistent flushes with uniform fruit bodies. |
| Testing for contamination | Before inoculating expensive grain jars, you can test spore syringes, liquid cultures, or tissue samples on agar first. Contamination shows up clearly on agar within days, saving you from wasting grain and time. |
| Cloning | Found a perfect mushroom? Take a tissue sample from the inside of the stem, place it on agar, and clone that exact genetics. Every fruit from that clone will have the same traits. |
| Long-term storage | Colonised agar plates sealed with parafilm and stored in the fridge remain viable for 2 to 6 months. This lets you bank genetics without maintaining active liquid cultures. |
| Faster colonisation | Transfers from agar to grain or liquid culture use actively growing mycelium, which colonises significantly faster than multispore inoculation. |
Equipment Needed
| Item | Notes |
|---|---|
| Pressure cooker | Must reach 15 PSI. Stovetop models or Instant Pot work. Essential for sterilising agar medium. |
| Petri dishes or small mason jars | Plastic or glass petri dishes (90 mm or 100 mm). Alternatively, use small 125 ml (4 oz) mason jars — they are reusable and easier to sterilise. Condiment cups (no-pour) also work. |
| Parafilm | Stretchy wax film used to seal plates. Allows gas exchange while preventing contamination. Micropore tape works as a substitute but is less effective. |
| Scalpel or inoculation needle | A #10 or #11 scalpel blade on a handle, or an inoculation loop/needle. Used for cutting agar transfers and tissue samples. |
| Still Air Box (SAB) | A large clear plastic tub with two arm holes. All agar work must be done inside a SAB or in front of a laminar flow hood to minimise airborne contamination. |
| Alcohol lamp or lighter | For flame-sterilising your scalpel between transfers. An alcohol lamp provides a steadier flame. |
| Isopropyl alcohol (70%) | For wiping down surfaces, hands, and equipment. 70% is more effective than higher concentrations for killing microbes. |
| Measuring scale | Accurate to 1 g for measuring agar powder and nutrients. |
Tip: If you are new to agar work, start with no-pour agar in small mason jars. You mix the agar medium directly in the jar, sterilise it, and let it set at an angle. No pouring required, which dramatically reduces contamination risk.
Base Agar Recipes
All recipes below make approximately 1 litre of agar medium, enough for 40–50 petri dishes or 20–25 small mason jars. Scale proportionally for smaller batches.
About agar powder: The "agar powder" used in all these recipes is the same product sold as agar agar in grocery stores and health food shops. It is a plant-based gelling agent derived from seaweed. You can buy it from supermarkets (Asian food aisle), health food stores, or online mycology suppliers. Make sure you get powder form, not flakes or strips — the powder dissolves much more evenly. Food-grade agar agar works perfectly for mushroom cultivation.
MEA — Malt Extract Agar
| Ingredient | Amount |
|---|---|
| Light malt extract | 20 g |
| Agar powder | 20 g |
| Water | 1 L |
Best all-round agar recipe. MEA is the standard choice for most mushroom species. It provides a balanced nutrient profile that supports strong mycelial growth without being overly rich. This is the recipe to start with if you are new to agar work. Malt extract is readily available from homebrew suppliers.
PDA — Potato Dextrose Agar
| Ingredient | Amount |
|---|---|
| Diced potato (peeled) | 200 g |
| Dextrose (glucose) | 20 g |
| Agar powder | 20 g |
| Water | Top up to 1 L |
Good for wood-loving species. Boil the diced potato in 1 L of water for 20 minutes until soft. Strain through a fine mesh or cheesecloth — discard the potato pieces and keep only the starchy liquid. Add water back up to 1 L, then stir in the dextrose and agar powder. The starch content from the potato provides complex carbohydrates that wood-loving species like oysters, shiitake, and turkey tail respond well to.
PDYA — Potato Dextrose Yeast Agar
| Ingredient | Amount |
|---|---|
| Diced potato (peeled) | 200 g |
| Dextrose (glucose) | 20 g |
| Nutritional yeast | 2 g |
| Agar powder | 20 g |
| Water | Top up to 1 L |
Accelerates growth for slow colonisers. Identical preparation to PDA, but with the addition of nutritional yeast. The yeast provides B vitamins and amino acids that boost mycelial growth rate. Particularly useful for slower-growing species like Lion's Mane that benefit from extra nutrition.
DFA — Dog Food Agar
| Ingredient | Amount |
|---|---|
| Dry dog food (blended to powder) | 10 g |
| Agar powder | 20 g |
| Water | 1 L |
Cheap alternative. Blend dry dog food (any basic brand) into a fine powder and mix with water and agar. The dog food provides proteins, fats, and carbohydrates in a single ingredient. The resulting agar will be opaque and brownish. It works surprisingly well for most species, though the plates are harder to read visually due to the dark colour. This is a good option if you want to practise agar technique without spending money on specialty ingredients.
Tip: For DFA, choose a basic dry dog food without artificial colours or strong flavourings. Grain-based varieties tend to work better than grain-free formulas.
SDAY — Sabouraud Dextrose Agar + Yeast
| Ingredient | Amount |
|---|---|
| Dextrose (glucose) | 40 g |
| Peptone | 10 g |
| Nutritional yeast | 2 g |
| Agar powder | 20 g |
| Water | 1 L |
Specialist recipe for Cordyceps. Cordyceps species have unique nutritional requirements that standard mushroom agar recipes do not satisfy. The high dextrose content and peptone (a protein digest) mimic the insect-based nutrition that Cordyceps naturally feeds on. This is the standard medium used in commercial Cordyceps cultivation.
Strain-Specific Agar Recommendations
While most species will grow on most agar types, choosing the right medium for your strain gives noticeably faster colonisation and healthier mycelium. Use this table as a quick reference.
| Strain | Recommended Agar | Notes |
|---|---|---|
| Cubensis (Golden Teacher, B+, PE, Thai, etc.) | MEA | Fastest growth on MEA. Cubensis are not fussy — MEA is the clear winner for speed and consistency. PDA also works but is slightly slower. |
| Oysters (Pink, Blue, King) | PDA or PDYA | Oysters prefer starch-based media. The potato starch in PDA supports their aggressive growth habit. PDYA with added yeast gives even faster results, especially for King Oyster which is slower than other oyster varieties. |
| Lion's Mane | PDYA (with extra yeast — 4 g) | Lion's Mane is a notoriously slow grower on agar. Double the yeast in PDYA to 4 g per litre. The extra B vitamins and amino acids make a noticeable difference in growth speed. Expect 10–14 days for visible colonisation even with optimal media. |
| Shiitake | PDA or MEA | Both work well for Shiitake. PDA gives slightly better results due to the starch content, but MEA is perfectly adequate. Moderate growth speed. |
| Reishi | MEA | Reishi grows slowly on agar regardless of medium. MEA provides sufficient nutrition without being overly rich. Expect thick, lacquer-like mycelium that may produce a reddish-brown colour as it matures. |
| Turkey Tail | PDA | Turkey Tail produces thin, spreading mycelium that colonises quickly on starch-based media. PDA is the standard choice. Growth is typically faster than most wood-loving species. |
| Cordyceps | SDAY | Cordyceps have specific nutritional needs that standard agar recipes cannot meet. SDAY is required — do not substitute MEA or PDA. Growth is slow and the mycelium appears pale orange to white. |
| Wavy Caps | PDA (with wood extract) | Prepare PDA but replace the plain water with wood extract: boil 50–100 g of hardwood chips (oak, beech, or alder) in 1 L of water for 30 minutes, strain, then use this wood-infused water to make PDA. The lignin and tannins from the wood extract mimic their natural substrate. |
Warning: Do not use softwood (pine, cedar, spruce) for wood extract. Softwoods contain antimicrobial resins and terpenes that inhibit mycelial growth. Always use hardwoods.
Step by Step: Making Agar Plates
- Mix your chosen recipe.Combine all ingredients in a heat-safe container or flask. Stir well. The agar powder will not fully dissolve until heated — this is normal. It will look cloudy and grainy.
- Distribute into containers.Pour approximately 15–20 ml into each petri dish, or 20–30 ml into each small mason jar. If using mason jars, loosely screw on the lids. If using petri dishes, stack them and wrap the stack in aluminium foil.Tip: For the no-pour method, mix the recipe directly in each small mason jar (scale the recipe proportionally), loosely cap, and pressure cook. After sterilisation, lay the jars on their side at an angle to create a slanted agar surface. No pouring required.
- Sterilise. Place containers on a trivet inside your pressure cooker. Do not stack petri dishes more than 4 high. Bring to 15 PSI and hold for 45 minutes. Agar requires a longer sterilisation time than liquid culture because it is thicker and heat penetrates more slowly.
- Cool and pour (if using petri dishes).After the pressure cooker depressurises naturally, carefully remove the flask or container of liquid agar. Let it cool until you can comfortably hold the container (roughly 50–55°C) — hot enough to remain liquid but cool enough that it will not crack plastic dishes or create excessive condensation. Working inside your SAB, pour approximately 15–20 ml into each petri dish. Work quickly and smoothly. Replace the petri dish lid immediately after pouring.
- Let the agar set.Leave the plates undisturbed on a level surface for 30–60 minutes. The agar will solidify into a firm, slightly translucent gel. If you see excessive condensation on the lids, you can briefly crack them open inside the SAB to let moisture escape, then reseal.
- Seal with parafilm. Wrap a strip of parafilm around the seam of each plate, stretching it as you go. Parafilm sticks to itself when stretched and provides a contamination-resistant seal while still allowing gas exchange.
- Store or use. Unused plates can be stored upside down (to prevent condensation dripping onto the agar surface) in a clean, sealed container at room temperature for up to 2 weeks, or in the fridge for up to 2 months.
Danger: Agar must be sterilised at 15 PSI for a full 45 minutes. Unlike grain or liquid culture, agar's thick consistency means heat takes longer to penetrate. Under-sterilised agar will bloom with bacterial and mould contamination within days, wasting all your plates.
Inoculating Agar
There are three main ways to put mycelium onto agar plates. Each has its use case.
From a Spore Syringe
- Working inside your SAB, shake the spore syringe well to distribute spores.
- Flame the needle until red hot, then let it cool for 10 seconds.
- Crack open the petri dish lid just enough to access the agar surface.
- Drop 1–2 drops of spore solution onto the agar. You can make a zigzag line across the plate for better distribution.
- Close the lid immediately and seal with parafilm.
Spore inoculation produces the most genetic variation. You will see multiple germination points with different growth patterns. This is where sector isolation (covered below) becomes essential.
From Liquid Culture
- Working inside your SAB, draw up a small amount of LC into a sterile syringe.
- Flame the needle and let it cool.
- Drop 1–2 drops onto the centre of the agar plate.
- Seal with parafilm.
LC-to-agar inoculation is primarily used for testing liquid culture for contamination before using it on grain. You should see clean, uniform mycelial growth from the drop points within 3–5 days.
From a Tissue Clone
- Select a healthy, mature mushroom with the traits you want to preserve (size, shape, cluster formation, speed).
- Working inside your SAB, tear the mushroom stem in half lengthwise with clean hands. This exposes the sterile inner tissue.
- Using a flame-sterilised scalpel, cut a small piece of tissue (3–5 mm) from the very centre of the exposed inner stem. Do not touch the outer surface — it is covered in contaminants.
- Quickly open the petri dish lid, place the tissue piece on the agar surface, and reseal.
- Seal with parafilm.
Tissue cloning is the most powerful technique because it captures the exact genetics of a proven performer. The mycelium will grow outward from the tissue sample in a circular pattern within 3–7 days.
Tip: When cloning, always take tissue from the inner stem, never from the cap or outer surface. The inside of the stem is the cleanest tissue on the mushroom. Work quickly — every second the plate is open is an opportunity for contamination.
Reading Agar Plates
Learning to read agar plates is a critical skill. Here is what to look for.
Healthy Mycelium
| Growth Type | Appearance | What It Means |
|---|---|---|
| Rhizomorphic | Thick, ropy strands radiating outward like roots or lightning bolts | Strong, aggressive genetics. This is what you want to isolate and transfer. Rhizomorphic growth typically produces faster colonisation and better yields. |
| Tomentose | Fluffy, cotton-like, uniform white growth | Healthy but less aggressive. Common in multispore cultures. Can still produce good results but is generally slower to colonise. |
| Sectoring | Distinct pie-slice shaped sectors with different growth patterns on the same plate | Multiple genetics competing. Each sector is a different genetic individual from the multispore inoculation. This is normal and expected from spore syringes. |
Contamination Signs
| Contaminant | Appearance | Action |
|---|---|---|
| Trichoderma (trich) | Starts white, then turns bright green as it sporulates. Fast-growing, aggressive. | Discard the plate immediately. Do not open it — green trich spores spread rapidly through the air and can contaminate your workspace. |
| Bacterial contamination | Slimy, wet-looking patches. May be white, yellow, or orange. Often has a sour or unpleasant smell. | If the mycelium has not reached the bacteria, you may be able to transfer a clean piece of mycelium from the opposite side of the plate. If the bacteria is widespread, discard. |
| Black mould (Aspergillus) | Dark black or dark green spots, powdery texture. | Discard immediately. Do not open. |
| Cobweb mould | Extremely wispy, grey, fast-growing. Much thinner and greyer than tomentose mycelium. | Can sometimes be outrun by aggressive mycelium. A small amount of hydrogen peroxide (3%) dropped on the cobweb can kill it while leaving mycelium unharmed. Transfer clean mycelium away from the affected area. |
| Yeast contamination | Shiny, raised, cream-coloured spots or streaks. Looks wet and smooth. | Transfer clean mycelium away from the yeast. If widespread, discard. |
Danger: Never open a plate with visible green or black mould outside of a SAB. Mould spores are microscopic and become airborne instantly. Opening a contaminated plate in your growing area can spread spores that persist for weeks and contaminate future work.
Transfers and Sector Isolation
Transfers are the process of moving a small piece of mycelium from one agar plate to a fresh plate. This is how you isolate clean, strong genetics from a multispore culture.
Why Transfer?
- Clean up contaminated plates: If one side of a plate has contamination but the other has clean mycelium, you can transfer the clean mycelium to a fresh plate and leave the contamination behind.
- Isolate genetics: From a multispore plate showing sectors, you select the strongest sector (usually the most rhizomorphic growth) and transfer it to a fresh plate. Repeat 2–3 times until you have a plate with uniform, monoculture growth.
- Rejuvenate old cultures: Mycelium on agar loses vigour over time. Transferring the leading edge to fresh media revitalises it.
How to Transfer
- Prepare fresh agar plates in advance. You need them ready and sterile before starting transfers.
- Working inside your SAB, flame-sterilise your scalpel until the blade is red hot. Let it cool for 10–15 seconds.
- Open the source plate and cut a small wedge (approximately 5–10 mm) from the leading edge of the mycelium — the outermost growing front. This is where the youngest, most vigorous mycelium is.
- Quickly transfer the wedge to the centre of a fresh plate. Place the mycelium side down (the side that was touching the agar) onto the new plate.
- Seal both plates immediately with parafilm.
- Label with the date, strain name, and transfer number (T1, T2, T3, etc.).
Tip: Always take from the leading edge, never from the centre. The leading edge has the most actively dividing cells and is the furthest point from any contamination at the inoculation point. This is the golden rule of agar transfers.
Transfer Timeline for Isolation
| Transfer | Purpose | What to Look For |
|---|---|---|
| T1 (first transfer) | Move away from contamination or select a sector | Choose the strongest-looking sector from your original plate. You may still see multiple growth patterns on T1. |
| T2 (second transfer) | Further isolation | Growth should be more uniform. Select the leading edge of the most vigorous growth. Contamination should be absent. |
| T3 (third transfer) | Final isolation | You should have a completely uniform plate — single growth pattern, no sectors, no contamination. This is a monoculture ready for use. |
Most cultivators achieve clean, isolated genetics in 2–3 transfers. Beyond T4 or T5, there is a risk of senescence — the mycelium may lose vigour from too many transfers. If you are not clean by T3, your source culture may be too heavily contaminated to salvage.
Warning: Always re-flame your scalpel between every transfer. Touching a contaminated plate and then a clean plate without sterilising will spread contamination to every plate in your session.
Storage
| Method | Temperature | Shelf Life | Notes |
|---|---|---|---|
| Sealed plates at room temperature | 20–25°C (68–77°F) | 2–4 weeks | Mycelium will continue to grow and consume nutrients. Only suitable for short-term holding before use. |
| Sealed plates in fridge | 2–6°C (36–43°F) | 2–6 months | Cold slows mycelial metabolism to near-dormancy. Seal well with parafilm to prevent drying out. Store in a sealed container or zip-lock bag for extra protection. |
| Slants in small tubes (fridge) | 2–6°C (36–43°F) | 6–12 months | Agar slants in test tubes or small vials have less surface area exposed to air, reducing desiccation. Best method for long-term storage. |
Tip: When taking a culture out of cold storage, let it warm to room temperature for 24 hours before transferring to fresh media. The mycelium needs time to “wake up” from dormancy. Growth may be slow for the first few days but will return to normal speed as the culture reactivates.
Quick Reference: Recipe Comparison
| Recipe | Cost | Difficulty | Best For | Readability |
|---|---|---|---|---|
| MEA | Low | Easy | Cubensis, Reishi, general use | Excellent — clear, light-coloured agar |
| PDA | Very low | Moderate (potato prep) | Oysters, Shiitake, Turkey Tail, wood-lovers | Good — slightly cloudy |
| PDYA | Very low | Moderate (potato prep) | Lion's Mane, slow growers, oysters | Good — slightly cloudy |
| DFA | Very low | Easy | Practice, budget grows | Poor — dark and opaque |
| SDAY | Moderate | Easy (but needs peptone) | Cordyceps (required) | Good — clear, light-coloured |