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An Experiment in Growing a Bioluminescent Mushroom

Follow me as I experiment with growing the bioluminescent mushroom Panellus stipticus (aka “The Bitter Oyster”) from my home lab.

Last Updated on June 16, 2020

As a new member of the Los Angeles Mycological Society I was excited when I saw an email hit my inbox with an invitation to attend a presentation on the topic of “glow-in-the-dark”, or bioluminescent, mushrooms by Assistant Professor Brian A. Perry at Cal State East Bay.

Brain’s Presentation, dubbed Illuminating Fungi – The Science of Fungal Bioluminescense, walked us through the fascinating world of glowing mushrooms around the world and the current phylogenetic research he and his team are working on to better understand how luminescent fungi evolved this incredible ability independent of one-another, as well as any clues that might reconcile any taxonomic uncertainties.

He also gave an introduction to luciferase and luciferine, the enzymes and compounds that give rise to the bioluminescent reaction. I’ll be diving much deeper into these as I ramp up my DNA research in my home lab.

At the end of the presentation, I had the opportunity to ask Brian which species of luminescent mushroom would he recommend for home cultivation. His recommendation was to have a go at Mycena Chlorophos and ensure there is adequate caffeic acid available for the fungi to facilitate the enzymatic process of luminescence.

Up until that point, I had never heard of caffeic acid before. After doing a little research, I discovered that it’s a naturally occurring organic compound that naturally exists in all plants. Interestingly enough, it’s also used quite a bit in a cosmetics. If you search eBay for caffeic acid, you’ll see sellers marketing it as such.

Unfortunately, getting my hands on a culture of any Mycena fungi has proven to be difficult. I did; however, find an alternative that I’m moving forward with: Panellus stipticus aka The Bitter Oyster.

Determined to grow a glowing mushroom and capture some cool photos, I have decided to use this experiment as an opportunity to learn a variety of new mycological skills. Let’s dive into it.

Goals of the Experiment

This experiment presents an opportunity to learn and try many new tools, techniques and mediums which have been on my to-do list for some time. Although my main goal is to successfully cultivate and capture a bioluminescent mushroom, I’d like to take advantage of this project to learn some new skills that will needed for some of my future plans.

Here are some of the goals I put together for this experiment:

  • Replicate results from published research on the optimal growing conditions for maximizing luminosity of Panellus stipticus
  • Prepare and inoculate PG (potato glucose) liquid culture
  • Test my new custom lids with self-healing injection ports
  • Test which jar labels survive the pressure-cooker sterilization
  • Calibrate my cheap pH meter found on Amazon
  • Compare the luminosity of two strains across varying pH in liquid culture and in fruiting bodies
Panellus stipticus LC inoculation
Getting ready to inoculate the prepared LC jars

The Research

I am using two published research papers for this experiment:

1. Prasher, i.b. (2012). Prasher IB, Chandel VC and Ahluwalia AS. Influence of culture conditions on mycelial growth and luminescence of Panellus stipticus (bull.) P. Karst. Journal of research in Biology (2012) 3: 152 – 159. Journal of research in Biology. 3. 152-159.

2. Se, Medvedeva & Ks, Artemenko & Aa, Krivosheenko & Ag, Rusinova & Ek, Rodicheva & Ap, Puzyr & Bondar, Vladimir. (2014). Growth and light emission of luminous basidiomycetes cultivated on solid media and in submerged culture. Mycosphere. 5. 565−577. 10.5943/mycosphere/5/4/9.

I found these two publications by searching Google Scholar for Panellus stipticus and finding the most recent publications which were focused on assessing the affects of growing conditions on luminosity. Said differently: I wanted a paper that would tell me how to grow my cultures to maximize the brightness of the fungi.

The main findings I’m using to determine how I organize this experiment are as follows:

The greatest light emission was obtained when Panellus stipticus mycelia were grown on PGA and SabA (Fig. 2).

Panellus stipticus mycelium growing on all of the media exhibited long-term luminescence (more than 40 days) (Fig. 2, 4). Although the luminescence differed in the time of occurrence and intensity, its maximum was reached at certain times depending on the composition of the culture medium: after 4–5 days of cultivation on FA, after 9– 11 days of cultivation on PSA, after 11–14 days on PGA, after 10–11 days of cultivation on MEA, after 13–15 days on SabA and YMA. As noted above, Panellus stipticus mycelia did not reveal any considerable differences in growth of colonies on media with different compositions.

Se, Medvedeva & Ks, Artemenko & Aa, Krivosheenko & Ag, Rusinova & Ek, Rodicheva & Ap, Puzyr & Bondar, Vladimir. (2014)

Temperature and pH affect growth and bioluminescence to a great extent. Glucose-peptone medium has been found to be the best for optimum mycelial growth as well as luminescence. The fungus exhibits luminescence at 20-24°C. The maximum mycelia dry weight (mg/25ml of the basal media) and luminescence observed at pH 4.0. The fungus exhibits luminescence after eight days of incubation at 24°C and pH 4.0, whereas it intensified to maximum after 13 days of incubation (pH 4.0 and temperature 24°C).

Prasher, i.b. (2012). Prasher IB, Chandel VC and Ahluwalia AS. (2012)

I was originally only going to run this experiment with liquid cultures but after re-reading the papers, it looks like I’m going to need to grow out these species on solid agar mediums in order to maximize brightness. The solid mediums showed a significantly higher level of luminosity compared to their liquid culture counterparts.

Materials and Methods

For the experiment, I’m growing two strains of Panellus stipticus.

  1. Panellus Stipticus ME-P8 from MushroomEmporium.com 4/28 (Shipped from Maine)
  2. Panellus Stipticus from fungi_supply on eBay 4/24 (Shipped from Canada)

Here are the general materials and methods used to prepare the liquid cultures:

  • Boil 2 large peeled Russett potatoes on medium for an hour
  • Prepare 2L, 1L batch-wise, liquid culture (LC) using 200g/L strained potato broth and 20g/L Karo Dark Corn Syrup
  • Prepare 250mL Potato Glucose LC for 8 jars
  • Pressure-cooker sterilize culture jars for 20 mins at 15 PSI
  • Inoculate each jar with 1mL LC culture syringe (2 strains * 4 jars)
  • Store jars at room temp (my closet) and allow to incubate in the dark
Preparing Potato for PG Liquid Culture
Liquid Culture Prep
Experiment Setup

Updated 5/2/2020

After re-reading the aforementioned studies, it occurred to me that liquid cultures yielded the lease amount of luminescence over solid growth mediums. In order to improve my chances of capturing the luminescence with the naked eye, I have added two grain and two MEA agar cultures into the mix.

The plates and grain jars were innoculated from a selection meeting the following criteria:
* Jars and plates would represent both A & B strains
* LCs would have pH closests to 3-4 which the research showed optimized luminosity
* I would take the LCs which shows the most aggressive growth so far

Accordingly, here is a photo of the new additions to the experiment with the source liquid cultures. Note the growth that is visible at the bottom of the jars.

The grain jars contain winter rye berries PC for 90 mins @ 15PSI. The jars were inoculated with 5mL and the plates with 1mL of LC via syringe.

Additional Cultures of Experiment
Solid and Grain Media Cultures Added to the Experiment on 6/2/2020.

Results

I am working on finding a better way to track and share the experiment results. For now, here’s an initial overlay of the experiment variants.

Come back here to find updated results and *hopefully* some really cool glowing mushroom photos as the experiment progresses.

Updates
Be sure to check out the following for updates to this experiment, including cool photos and more:

Bioluminescent Mushroom Grow (Update #1)

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