This document provides instructions for cultivating mushroom spawn from start to finish. It begins by defining mushroom spawn as any substance inoculated with fungal mycelium to promote fungal growth. It then discusses the importance of maintaining clean spawn to avoid contamination. The document outlines the materials needed, such as an autoclave, HEPA hood, petri dishes, jars, and filters. It provides step-by-step instructions for transferring clean cultures from petri dishes to jars of spawn substrate like grains. The goal is to generate healthy, contamination-free mycelium ready to inoculate fruiting substrates.

1. +
+
A Guide to
Cultivating
Mushroom
Spawn
Mushroom Spawn: any substance that has been
inoculated with fungus mycelium to promote
growth of the vegetative stage of fungi life cycle.
Scientists, farmers, hobbyists, and people of all walks of life are exploring the realms of
fungi cultivation. Whether for research, food production, or basic interest in the field,
the most important aspect of producing your own fungi is maintaining clean spawn.

2. 2
Some Reasons to Cultivate Fungi!
Humanity has always shared a strong tie with fungi, whether producing alcoholic beverages,
consumption in diets, uses in medicines, or guides to spiritual enlightenment. Regardless of reasoning,
the art of mushroom cultivation is a satisfying adventure for beginners and veterans alike. Here are a
few examples of fields of interest in mushroom cultivation:
Culture/Health Benefits
 The Chinese culture has always revered the Ling Zhi mushroom (Ganoderma lucidum) for its
good luck traits in supporting life and longevity; modern science has gone on to assure the
benefits of Reishi consumption in cancer treatments, diabetes prevention, and neuroprotection
relating to dopamine levels in the brain [1,2,3].
Beauty
 The turkey tail mushroom (Trametes versicolor), admired by mycophiles for its beautiful
appearance, exhibits anti-cancerous traits by the simple consumption of the raw fruiting body
[4].
Diet
 The all-common oyster mushroom (Pleurotus ostreatus) is enjoyed worldwide for its delectable
taste, and ease in cultivation. Modern science is currently investigating health benefits of
complex polysaccharide chains present in fruiting bodies [5].
Top left: rare Pink Oyster mushroom fruiting on tree trunk
(http://mycosource.com/wp-content/uploads/2012/08/Pink-Oyster.jpg)
Top Right: Exquisite Ganoderma lucidumflashing colors
(http://www.npnutra.com/assets/images/reishi-mushroom-2.jpg)
Bottom left: Turkey Tail mushroom fruiting on tree trunk
(http://i.ytimg.com/vi/UN9RcvrUw64/hqdefault.jpg)

3. 3
What is mushroom spawn and why is it so
essential to growing mushrooms?
Similar to plants, fungi must first
establish a vegetative body to prepare
for fruiting. Fungi achieve this by
producing mycelium (pictured on the
bottom right); mycelium is an
interconnected, branching, web-like
network of cells that grow in the soil
or over a substrate. Substrates are the
growing medium that mycelium
colonize and enzymatically break down
for access to simple sugars that spur
growth. In growing out this young
mycelium, and letting it grow/colonize
over substrate lays the essence of
mushroom spawn. After colonization of
substrate, the mycelium will have
established vegetative growth ready
inoculate more fresh substrate.
To achieve this vegetative growth,
spawn substrate is most commonly
grown on sawdust, or grain depending
on the type of fungi being cultivated.
For example, lion’s mane (Hericium
erinaceus) prefers an oak sawdust
substrate, compared to oyster
mushrooms whose mycelium prefers
that of grain.
After colonization of spawn substrate is
completed, the transfer to the fruiting
substrate is necessary.
Upon complete colonization of a spawn
substrate, the spawn is transferred to
fruiting substrates. When a transfer of
mycelium from one substrate to another
occurs, this is known as an inoculation.
Transfers and inoculations are forms of
propagation; or increasing the number of
fungi cultures. Most fungi prefer materials
such as straw, sawdust, cardboard, paper,
woodchips, or even compost for fruiting
substrates.
Top: Grain jar uncolonized, a
simple mass of individual grain
kernels (spawn substrate)(Voss
Picture)
Bottom: electron microscope of
mycelium (www.indiegogo.com)
While generating Mushroom spawn, the
primary goal is to establish healthy,
contamination free mycelium, which is
ready to inoculate. Spawn transfers
always lead to more vegetative mycelium,
but once applied to the correct substrate,
fruiting will ensue. Quality spawn is of
the utmost importance while attempting
to fruit mushrooms.
First, Some Basic Myco-Vocabulary
Simplified Substrate
Spawn Substrate: better triggers growth
of vegetative mycelium
Fruiting Substrate: better triggers
growth of mycelium leading to healthy
fruiting bodies
While this brochure will touch slightly on mushroom fruiting techniques, the primary
goal is to emphasize mushroom spawn cultivation.
The discussed techniques are in regard to cultivation techniques in a sterile lab
production scenario outside of nature.

4. 4
What Materials Do I Need to
Begin Spawn Production?
Autoclave or Pressure Cooker
 Allows one to sterilize media of contaminants before
inoculation
HEPA Fume Hood
 HEPA: High efficiency particulate arrestance
 This allows one to work in sterile conditions while
transferring mycelium from growing media to new
growing media.
 Room should be kept sanitary as much as possible
Sterilized Petri Dishes
 There are many agar, malt, yeast combinations that
can be combined with other chemicals to maximize
selection for fungi while avoiding contaminants
Sterile tissue technique tools
 Scalpel, Bunsen burner, tweezers
 Ethanol or other sterilizing agent
Standard sized Mason Jars (Bottles)
 Cut two small holes in top for air flow
Microporous filters
 Placed between lid and jar to allow air flow that
withstands contaminants
Mushroom culture or live mushroom specimen to be
cloned
So if fungi occur naturally outdoors in diverse
ecosystems, then why do I have to be so careful to avoid
bacteria and other fungi in my mushroom cultivation
practice?
*The mushroom cultivator is not trying to replicate an
ecosystem in all of its complexities, but instead replicate
an environment for one specific fungus to flourish. If
even a single contaminant jeopardizes spawn or
substrate, the mycelium will then have competition over
the small amounts of food. This competition of the
microenvironment leads to weaker mycelium leading to
less hardy, or even rotten mushrooms.
Note:
The most essential aspect of producing mushroom spawn is
avoiding contamination at all costs. One should be very
thoughtful to keep a watchful eye for pests, and unclean
looking bacteria in your petri dishes, jars, bags, or substrates
of any caliber. Contaminated spawn will transfer
contaminants. Once contaminated, your spawn is garbage.

What’s the Big Deal With
Contamination?
HEPA Fume Hood
Diagram showing
function
(http://www.baylor.edu/
content/imglib/1/9/4/6/1
94643.jpg)
Standard autoclave
used to pressure
cook substrates of
contaminants
(http://www.zirbus.co
m/autoclaves-
laboratory.png)
Right: Mason jar infected with
Bacillus sp.
Left: cap infected (unknown)
(www.shroomery.org/)

5. 5
+First things First: Use clean culture or isolate from
fungus sample
1. Utilizing a mycelial culture already in a test tube or
petri dish, using a sterile tool, transfer a small amount
of mycelium (white biomass in the picture above) to a
new petri dish.
2. Wrap edges of Petri dish with Parafilm to seal the
microenvironment.
3. Allow enough time to for the fungi to colonize the
whole dish. (1-2 weeks)
4. Watch for contamination that often manifests in an
abstract color, fast growth rate, or shiny texture.
As pictured above, the petri plate of “Po Wellington P:6” is
sterile culture indicated by uniform white mycelium. If
your culture does contaminate, one must re-transfer into a
new plate. Contamination this early in the process will
certainly wreak havoc further down the line of transferring
your mycelium to different media. Once your culture is
colonized and clean, it is time to transfer to a new
medium. [6]
For the more ambitious individuals, samples can be
isolated directly from specimen of fungi in the wild,
from a friend’s, or from one that is to be cooked for
dinner.
1. Instead of transferring culture to culture, use a
scalpel or another tool to grab a piece of white
mycelium from the stipe (stem) and place in the
petri dish. If the stipe is not offering prominent
mycelium, samples can be taken from the spore-
heavy gills on the bottom side of the mushroom
cap. The picture above features a diagram of
grabbing mycelium from the top gills, or the
bottom stipe.
2. Ample time is given for mycelia to colonize
contamination free.
3. The isolation should be transferred to another petri
dish in the same format described on the left side of
this page. This excess step is to make sure no
unseen contaminants are persisting.
Clean Culture Transfer Mushroom Isolation

6. 6
+Plate to Jar
Transfer
After the Petri dish is completely
colonized, transfers to jars full of
spawn substrate will follow.
Again done under your HEPA
Fume Hood, like all inoculations of
new, sterilized substrate:
1. Simply cut the Petri plate into
wide chunks (some like to cut
the circular plate into 8 slices
like a pizza)
2. Drop the slices of mycelium
into your sterilized jar of
grain. Leave about 2 inches
free to allow mixing of grain
upon inoculation.
3. After the jar is propagated,
seal with microporous filter
between the jar top and lid.
4. Shake mycelium throughout
jar to evenly distribute
It should be noted that the
importance of moisture content of
your grain cannot be
overemphasized. Grain should be
soaked for 12+ hours before
autoclaving and inoculation. The
grain will soak up a large amount
of water necessary for mycelium
to grow.
After your jar is inoculated, precaution should be taken to shake the bottle in
a way to spread the slices evenly throughout the grain substrate. Instead of
allowing the mycelium to all be in one place, rearranging positions of
mycelium throughout the jar will lead to more points of inoculation and a
faster colonization throughout the whole. After one week, the jars will be near
fully colonized as shown in the above picture. After one and a half weeks the
jars will be ready for transfer once more. [7].
Post colonization, the grains will be fused together by the dense network of
mycelium laid throughout. Before transferring, the jar should be smacked
around and shaken up to break up the grains until they are freely moving
once again. This breaking of the mycelium creates a vast amount of
inoculation points on every mycelial piece that aids in faster colonization on
the next propagation step.
At this point you have now successfully made grain spawn! After colonization,
this mycelium has established vegetative tissue for some time, and is ready to
spread its spores via fruiting! If your attention to detail and contamination
has made it this far, there is good chance you’ll be fruiting mushrooms in no
time!
“The art of propagating mushrooms is primarily the art of transferring the
mycelium from one food source to another to keep it going as long as
possible.”
-Edmund Williams
Grain spawn jars partially colonized (Voss photo)

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Jar to Jar: More Propagation, or Fruiting!
Jar Transfers
Jar to Jar transfers can be utilized in generating more spawn, or to
begin the fruiting stage with Bottle Culture. Whether generating
more spawn, or beginning fruiting, the technique is the same, but
the difference lies in the substrate. Transferring bottled grain spawn
to a bottle with adequate substrate is a pretty simple process.
1. Under a HEPA fume hood, shake to break up the mycelium in
colonized jar
2. Pour broken up spawn into a new jar with preferred substrate
that has been sterilized. (Keep a little space at top so mixing
sealed jars is possible)
3. Seal jars with lid and microporous filter.
4. Shake to disperse spawn evenly.
One of the major benefits of bottled grain spawn is that it affords
the possibility of further propagation in large amounts if desired.
One bottle of completely colonized grain spawn can be used to
inoculate ten more bottles. Imagine having ten jars of colonized
grain spawn; it is very possible to make a quick jump to one
hundred bottles if every bottle inoculates ten more. Likewise,
fruiting substrates in jars can also be inoculated for high amounts of
mushrooms.
Jar to jar transfer can be used to spur Bottle Culture fruiting, or to
continue building up a spawn collection for future fruiting.
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The primary focus of Bottle Culture is to fruit mushrooms from the
bottle, not to make more spawn. Before attempting to fruit from the
jar, one should be aware that certain fungi fruit better from jars than
others, research is necessary. Instead of grain spawn throughout,
ample substrates such as sawdust are preferred for fruiting. After
sterilization of the bottle with the sawdust inside, grain spawn is often
used to inoculate bottles in a quick and efficient way. After
inoculation, the jars are shaken and given warm and dark
environmental conditions until the jar is colonized. Once colonized, the
lid is taken off the jar, humidity is raised, and light is introduced to
the culture to trigger fruiting.
Bottle culture often leads to uneven colonization throughout the jar
based on dispersion of inoculum grains. This usually leads to the top
of the jar colonizing before the bottom. This is beneficial in that the
oldest mycelium produces the fruiting bodies, but the mycelium at the
bottom of the jar is never able to fruit upon reaching maturity. This
method does not maximize fruiting, but offers a very quick, efficient,
and smooth operation [8].
If any contamination of the smallest degree is found in a jar, that jar
should be discarded. As continual propagation steps occur, the
contamination will manifest in larger amounts further down the line.
Fruiting: Bottle Culture

8. By: Samuel Voss vossx209@umn.edu
Final Considerations
Mycology is an ever-growing field that is undergoing
revolutionary cultivation techniques annually. Part of the
fun in caring for fungus is the creativity the cultivator can
explore in maximizing their product. Whether
experimenting with the wide array of different species,
altering substrate to see what the fungus prefers, altering
environmental conditions, or simply foraging in a forest,
there are many routes to taking up a myco-hobby for
enjoyment. Furthermore, after the mushroom growth
experiments are concluded, everything involved in the
equation will be compostable if organic compounds are
strictly used!
This brochure highlighted a very direct and distinct avenue
of mushroom grain spawn cultivation while also touching
on one fruiting technique, yet there are many more avenues
available in producing mushrooms including bag spawn
fruiting, outdoor patch inoculations, tree stump
inoculations, and straight log inoculations to name a few.
While there are many ways to cultivate fungus, quality
spawn is necessary for any growth of value to occur. Sterile
tissue technique to grain spawn is a tried and tested way of
producing highly efficient mycelium spawn in a format that
can maximize quality and quantity. By learning to control
contamination variables, the myco-cultivator is one step
closer to achieving success.
Top picture: bags of
ash sawdust before
being autoclaved,will
be inoculated with
oystermushrooms
after. (Voss Picture)
Picture of the author
holding PoHu Oyster
cluster (Pleurotus
ostreatus) and a large
Lion’s Mane fruiting
body (Hericium
erinaceus)
Endnotes
1. Liang, Z (2014). "Ganoderma lucidum polysaccharides target a Fas/Caspase dependent pathway to induce apoptosis in human colon
cancer cells". Asian Pacific Journal of Cancer Prevention 15 (9): 3981–3986.
2. Teng, BS (2012). "Hypoglycemic effect and mechanism of a proteoglycan from ganoderma lucidum on streptozotocin-induced type 2
diabetic rats.". Eur Rev Med Pharmacol Sci 16 (2): 166–75.
3. Zhang, R (2011). "Ganoderma lucidum Protects Dopaminergic Neuron Degeneration through Inhibition of Microglial Activation.".
Evid Based Complement Alternat Med. doi:10.1093/ecam/nep075
4. "Antimetastatic effects of PSK (Krestin), a protein-bound polysaccharide obtained from basidiomycetes: an overview". Cancer
Epidemiol. Biomarkers Prev. 4 (3): 275–81. 1995. PMID 7606203
5. Wasser, S.P., 2002. Medicinal mushrooms as asource of antitumor and immunomodulating polysaccharides (minireview).
Appl. Microbiol.Biotechnol., 60: 258-274.
6. Stamets, Paul. Growing Gourmet and Medicinal Mushrooms. Volume 3. Berkeley: Ten Speed Press, 2000. 94-96. Print.
7. Stamets, 121-128. Print.
8. Stamets, 191-193. Print.