Mushroom DNA Sequencing with Kyle Canan
Alisa Nappa
Today we sit down with the robin hood of mushroom DNA sequencing, Kyle Canan, founder of the Ohio Mushroom DNA Lab, which offers free mushroom DNA sequencing to the public. We chat about the process of DNA sequencing, finding new species, types and cost of equipment, how to set up your own lab, how to send in your own samples and more.
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TRANSCRIPT
Unknown Speaker 0:11
Welcome, welcome. You are listening to the mushroom revival podcast. I'm your host, Alex Dora. And we are absolutely obsessed with the wonderful, wacky world of mushrooms and fungi. We bring on guests and experts from all around the world to geek out with us and figure out what the heck is going on with these weird mysterious organisms that are fungi and mushrooms. So today, we have Kyle joining us from Ohio to talk about DNA sequencing and all the fun stuff that you can
Unknown Speaker 0:43
understand about the inner workings of fungi. So, Kyle, how you doing, man? Great. Thanks for having me.
Unknown Speaker 0:51
Yeah, so what do you what are you up to? In Ohio? What? What, what brought you to DNA sequencing? Yeah, cool. Um, so I'll just give a little background about myself quickly. So first and foremost. So Mike, naturalist and environmental environmentalist by heart.
Unknown Speaker 1:07
And I got into fungi several years ago, because of like medicinal reasons. At a young age, in my early like, you know, 20s, late teens, I was diagnosed with rheumatoid arthritis. And I was taking a few different types of drugs, you know, Western medicine style, and they were like injections, and it just didn't stick with me well, and I tried to get off the medication. And I did, and I was looking for all kinds of alternatives. And I kind of heard about, like Turkey Tail and being like, immune support and
Unknown Speaker 1:44
supporting like function. And I was like, wow, this is interesting. So it kind of just started to dig a little deeper on that. And I had the time I was unemployed. And I was kind of bored. And so mushrooms seems super interesting after after, like the Turkey Tail thing. And I was like, let me look more into this. And then I just kind of did. And
Unknown Speaker 2:05
one thing just led to another.
Unknown Speaker 2:08
I started using an app called I naturalist to identify my, you know, my mushroom observations that I would find and I think like Alan had recommended and some other folks
Unknown Speaker 2:20
and then just continued to study I quickly fell in the mycology community. He like,
Unknown Speaker 2:27
I found my my, my little niche, and I fit right in I quickly like became part of the local mushroom society here and just started doing everything it took to learn more about mushrooms every which way. And I was just fascinated. And then in 2021, I was on a winter tree hike. So in Ohio, we get some pretty crazy winters. It's like 20 degrees today, actually. So not many mushrooms out. In the winter months, that usually, I would say, like December, all the way up until the end of March. So I was on a hike. And today I was 21 all focused on trees, because I'm a big tree lover as well, like, I'm a county coordinator for the North Forest network. So I'm always trying to go to old growth forests. That's, that's my thing. Like I actually have a little like, uppity up when it comes to mushroom collecting because I won't like go to a place that's been cut. I'm just like, I'll go to old growth cuz I just know what happens there. So we're at old growth forests. And
Unknown Speaker 3:35
at the end of the hike, I was with a friend at the time, we were just chit chatting about trees, we look down and there's this pile of mushrooms alone. That instantly I was like, wow, this is crazy, because of the time of year. And then it turned out to be a philosophy mushroom, and further turned out to be a new philosophy mushroom. So
Unknown Speaker 3:59
at the time, I was just chatting it up with Alan Rockefeller. I sent him pictures. I'm like, Yo, you got to see this. We looked into the mushroom a little deeper from the morphology of the pictures. He's like, You got to send this to me. It's got to be sequenced.
Unknown Speaker 4:14
What is sequencing? Really? What is this? I no idea never really heard the terms allow. So it's enough Allen guess sequenced and got some results back saying it was a new philosophy.
Unknown Speaker 4:25
After that, everything kind of just spiraled in the good way in the Hawaiian live the founder of the wild mushroom DNA lab. So sorry, that was a little long, but that was that's the intro story. And I'm probably going to completely butcher this Latin name, but that was Salafi. cero Leo Ryza, Nam prov, right. Something like that. Philosophy cilia. Ryza. Nonprofit.
Unknown Speaker 4:51
It Latins tough struggle with it. So I get Sue Uriah Allen gave it the name at the time Marilyn
Unknown Speaker 5:00
Did you know he was the only one doing the DNA sequencing and other than another one of my colleagues, Stephen Russell, at the time, I wasn't aware of who Stephen was only knew Allen Allen to sequence it when had put the lat and on there. And yeah, actually, Alan, myself and a couple of my other team members are describing as we speak. So in 2024, in Sri Lanka, the nonverbal drop off. Sweet, amazing, and when can you tell me more about this species? Has there been any trip reports? You know, what, anything pretty special about it? Anyone had tried to grow it? You know? Yeah. Okay. So it is a word loving philosophy. So when we found that when I found it, it was instantly like, Whoa, this is philosophy of audio stereo out of because it is found in Ohio and the PA region, and you know, the Ohio River Valley basin. And that's kind of where it was, it was it was in the suburbs of Cincinnati sells like, well, this is
Unknown Speaker 5:57
old weights. And the date just seemed seemed different. And then after further examination of the morphology of the spores, we were able to quickly tell that there this isn't an ovoid. So what we know is it's a wood lover, it's growing in disturbed, mulch landscape areas.
Unknown Speaker 6:18
The Ohio location seems to be really popping.
Unknown Speaker 6:23
And it's only other have been found in one other location in Ohio confirmed their DNA. And it was at a truckstop also had malt. And you would think we'd be finding a lot more, but we're not the four first reporting states through Ohio, Indiana, Iowa, and Pennsylvania. And those all are confirmed through DNA. And then there's suspect in Michigan, unconfirmed. So will we also know is it is a winter philosophy, which is kind of just abnormal, right? What what months does it grow? Yeah, so it grows. It's been found in the months between November December.
Unknown Speaker 7:09
I could be a little longer than maybe late October, but I'm thinking it's only November, December, that helps your serotonin get through the winter.
Unknown Speaker 7:19
Definitely does. So I actually found in 2021, and I was one of the first discoverers of it, then, then I did not find it again in 2022. At all. I was actually out to Alan's house during the season. So I must have missed it. Then, in 2023, we had a great friend that lives down in Cincinnati area, she gave me a call a couple of weeks ago, so you need to get down here, it was probably more than a couple weeks ago, at this point it probably four or five. So I found it again. Last year, got some really good camera photos of it for Paul Stevens his book, and obviously the description that we're working on.
Unknown Speaker 7:57
And I'm sure there's quite a few guerrilla growers helping that
Unknown Speaker 8:03
something the growth of it, which, you know, happened with like, a lead, EI and ovoids is people are like, Oh, this is pretty easy to grow, like, you know, throw it in every wood chip pile around my area. Now, it's now that you can find them in a lot of places, which is exciting. It is for sure. So it's a continue answering some of your questions. Trip Reports. The only trip report I'm aware of and I think most of community is aware of is my trip report. So I would say they're moderate, moderately to highly active,
Unknown Speaker 8:35
you know, a little bit more than a microdose. And it was a pretty good experience.
Unknown Speaker 8:41
As far as growing, I'm not aware of anybody growing it yet. I do know a couple of people that are working on it.
Unknown Speaker 8:48
And then this species will also have HPLC reports and analysis done. So it'll be sent off to a chemist of Oregon. Jordan Jacobs in Jordan will run all those analysis and then we'll also be in the description pick. Three, I can't wait to hear what the levels of compounds if there's any new compounds. Yeah, that'd be him. What was your experience like compared to, you know, typical cubensis or, you know, other other species? I would say almost, it's all really the same to me. I do feel like it was stronger. So like I if I am going to dose myself it's either cubensis or ovoids because I find them often in Ohio in the spring that I can save up so I definitely would say the trip reminded me a lot more of like an overweight trip. So maybe it's just the potency of them stronger, probably. But super pleasant. I had a great time. Talk to one of my good buddies on my on my volunteer team and we chatted it up and had a good talk and actually worked on the paper. Someone said it was pretty cool. Sweet and would you say it's
Unknown Speaker 9:58
what? What level of
Unknown Speaker 10:00
You know, aggressiveness would you say that the mycelium has like taking over wood chips? Is it pretty aggressive? And obviously it's resilient, that it can survive and those low temperatures but considering it hasn't been found in many places, you know, what, is it pretty easy to grow? Or? That's a good question. I would like to say yes based off like the the the rise of morphic structures. The reason like Soulier rise was one of the names was because when
Unknown Speaker 10:32
I think how long was the first guy to put it on a guitar, it just like completely stained blue I don't know if you've seen why the egg or pictures but it is one of the most intense and blowin awesome just things in nature I've probably ever seen.
Unknown Speaker 10:46
So based off that and based off the you know, the observation myself all the, you know, the colonize wood chips, I would say so, however, I've tried to start my own here at home, and it's not doing so successful. So yeah, I don't know how Alan's got the magic touch. Probably, in some of these growers, you know, like these people that really just take a super serious I have mad props and respect for them, they're gonna figure it somebody will grill this thing. And what they do they get it all figured out vocally, they report back to me, give me an easy way. So I don't really have time. So that's awesome. Well, I'm, that's super exciting. I can't wait for
Unknown Speaker 11:26
the official name to be, you know, officially published and, you know, all this analysis to figure out what, what compounds are in there more people trying it out. And both growing and testing it. But yeah, those new species are popping up left and right. Oh, mostly, mostly, thanks to Alan, he's, he's done quite quite a lot of legwork in this philosophy field. So that's super exciting that, you know, yeah, citizen sciences, scientists are coming together and just figuring out mycology in a deeper way. So that that's awesome, man, for sure. I would definitely say,
Unknown Speaker 12:03
due to DNA sequencing, it's definitely
Unknown Speaker 12:07
adding to that and accelerating, I think, before DNA sequencing, which, before DNA sequencing would be like this, the 50s. But I'm saying before DNA sequencing of like fungi, you know, most people are probably, if it wasn't, if it was only just discovered in that time, say it was being found maybe in December, people weren't probably keen to call in it, you know, some late some late ovoids. And I think, right, what people were saying in the pictures, and same thing goes with the one of my friends.
Unknown Speaker 12:41
discoveries in Florida of philosophy, nivia tropicalis, now improv,
Unknown Speaker 12:46
same thing, it was also getting called ovoids. But with the DNA sequencing technologies, now we're able to kind of like separate the species, and we're able to tell it, there's, there's still a lot of work to be done. And now, the taxonomical region, in the realm of mycology itself, there's there's a lot of discoveries, we're uncovering all kinds of new species like lid, literally every month for sure. What is what is knov Prov stands for, and is that just tagged on to any species that are not officially described it in a published paper? I can't think of the actual Latin it's known inprove Vu or something like that. And it basically is just a temp code named Mom, that's, you know, waiting to be described. Cool. And so going into DNA analysis, which is kind of your bread and butter. Now, what can you kind of give us an overview of the Ohio mushroom DNA lab? What? What do you got going on? What are your goals? Yeah, sure. Um, so yeah, I kind of started traditional Sanger sequencing just for myself, right? I was after the slow spiels like, well, this is really cool. I need to better understand this. This philosophy is not the only mushroom I have collected, I've collected pretty much everything have countered. So like, I gotta get to the bottom of this. So I quickly got a quick was saying your technology. And then I was just as quick as I got equipped, I was quickly coming to the conclusion that, well, this was going to cost a lot for the amount that I just wanted, like it wasn't anything other than like, myself just literally wanting more barcodes out there for the public. Because once I realized, the whole philosophy, sillier Ryza situation, it answered in a lot of questions on might already out and then also opened up was like, a lot of opportunity and possibility. Like, wait a minute, if philosophy is not the only genius that's going to have like these cool new discoveries, it's going to be everywhere. It was saying your technology wasn't going to work.
Unknown Speaker 14:49
In 20 through 2022, I met Stephen Russell, Stephen Ross, who's kind of like my great mentor colleague. He's responsible for an
Unknown Speaker 15:00
Everything that I know about nanopore sequencing essentially
Unknown Speaker 15:05
metarhizium, which was a class that he was holding to learn nanopore sequencing. And I think I was like one of the only people out of the class to follow through, I was definitely destined that that was going to be it. I just needed to figure out a way to sequence a lot of mushrooms, all of my collection, and then and then some for somewhat affordable costs. So when I found out about nanopore and transition to nanopore, they use most of my own funds to kind of get started up. And I've always been like,
Unknown Speaker 15:39
you know, supporter and appreciative of grassroots grassroots movements. So it kind of just Yeah, reach out to the public, hey, this is what we're doing. This is this isn't my goal.
Unknown Speaker 15:51
You know, believe in me, send me some money. It'll work out I promise. And it kind of did. Just producing the first couple nanopore ruins and getting individuals results off of the specimens that they collected that year, or even say, like people had been saving stuff for years before, like, it's saving. Mushroom collections isn't anything new and mycology, that's been going on since the beginning of mycology, it's all having the accessibility to technology to analyze them is what's new. So people are able to get mushrooms that they've had saved, sequenced, and get the results. So that kind of just was the start of it. And I really wasn't sure where it was gonna go. I knew that I wanted to sequence a bunch of mushrooms. And that's really all I wanted to do, I didn't really need to prove to anybody or anything. I just wanted to get more mushrooms barcoded, so everyone can understand it. And they were really turned into what what Ohio mushroom DNA lab is known now. And it's taking specimens from anywhere and everywhere, putting them in a queue, sequencing them getting the barcodes adding the information and to as many public databases and resources as possible. So everything that we do is free to everybody as far from like beginning to end. So the sequencing free, we put it on I naturalist, we put it on GenBank. We make records and make permanent records on myco map. It's all accessible to the public making accounts here and there, all you got to do.
Unknown Speaker 17:27
And so yeah, now we're one of the leading labs in the country.
Unknown Speaker 17:32
We work hand in hand with the other two labs, there's, you know, probably only a handful of people that are DNA barcoding fungi on the nanopore device. So that's where we're at with it.
Unknown Speaker 17:46
So, we have a wide range of listeners that tune in to our episodes, and some people probably do DNA sequencing full time they fully understand the ins in ins and outs of it. And then probably a lot of people who have no idea what DNA sequencing means, or a barcode or what that is.
Unknown Speaker 18:06
For me, I'm I'm very unfamiliar, you know, I can dabble a little bit, but I've, I've never done it myself.
Unknown Speaker 18:14
You know, I know DNA looks like a double helix. And there's, you know,
Unknown Speaker 18:20
you get the long string of code of A T C, G, and you know, it's basically like a code that is the makeup of of your, of your being, you know, but can you kind of go through the step by step process of what happens when you do a DNA sequence of a mushroom. So say you find one in the wild, you bring it back home, then what? What Yeah, so after you get it home, you have to dehydrate the mushroom is least,
Unknown Speaker 18:53
probably the most recommended practice. Now you can do quick extractions, with, you know, freshman material and tissue. But basically, you want to do a drop. So I recommend everybody you try the mushroom, so you can dry the mushroom. Now you're dealing with a very dried crunchy mushroom, essentially, right?
Unknown Speaker 19:10
First step is the extraction process. And you have to extract the DNA from materials essentially. So without going into every step, you basically get a piece of mushroom tissue
Unknown Speaker 19:22
that's very small, almost smaller than you can really imagine. You can obviously still see it, however, it's a very small piece that goes into a very small plastic tube.
Unknown Speaker 19:33
And that is considered just like one reaction.
Unknown Speaker 19:37
And then with a couple of added chemicals, and some heat using a machine. The DNA is extracted from that tissue in its in solution, ie like water and some buffers. So that's like the first thing is extracting the DNA. So now you have extracted DNA and solution in little tubes. Now you have to amplify the DNA
Unknown Speaker 20:00
because we are only looking at one
Unknown Speaker 20:03
region, in the whole genome of fungi. So to look at the full genome of fungi, it costs significantly more. And it takes a lot more time, and is very helpful. But it's not necessarily needed for what we're kind of working towards our goals here home deal. Oh, so now you need to amplify, so you use tack polymerase, some primers, and there's whole scientific method on them called PCR. And by amplify, what do you mean? Yeah, so you're basically making multiple copies of this small gene region. That way, it's blowed up into a scale to where we can use technology like computers and stuff to read it.
Unknown Speaker 20:47
So you're amplifying it in a machine called the thermo cycler, that heats and cools, and it works at different times and as a little schedule and program. And that runs for like, two to three hours. And now your, your material is now considered to be PCR over, you know, that step is at least snow you have done extraction, you've now conducted PCR. So now what these little tubes are going to look like is like this green solution could be but mainly green, could be clear, too. So it's a solution again, that contains, you know, the DNA inside that you can't see with the naked eye. And it's billions of copies, essentially. So now a sequencing machine can now read that sequence and put it what we're literally saying in a sequence of those nucleotide acids, which are the at C's and G's, which are unique to and that's your barcode. And that is the barcode. Yes. So
Unknown Speaker 21:53
the sequence of the IDS region is considered our bark. Yeah.
Unknown Speaker 21:59
So basically, yeah, that's the first like three main steps. And now you're all down to analysis, and then need to run it through computers, and then do a bunch of manual work. So basically, then the computer, sorts everything out and aligns them and then basically puts it to the specimen, very, very complicated processes. And then a human has to basically blast search that information amongst and against other databases, and then basically kind of make a judgement based off that information on this beats. And so it's not 100% automated, there still has to be a lot of like, huge physical human work on the backend.
Unknown Speaker 22:48
So they basically copy and paste that into a bunch of databases and see what matches. Yeah, basically, yeah, the gem bank is like the the primary one, and that is accessible to everybody. And it's a government website, they use it for genes of everything, right. Like, there's human genes on there, there's everything.
Unknown Speaker 23:09
And then we have a more local database that Stephen Russell created, and it's called Michroma. So we basically match that information against these other databases, and then a judgment call has to be made.
Unknown Speaker 23:24
You know, obviously, still, following the rules of mycology and taxonomy. It you know, there could be some very more complicated and some very easy, it just kind of depends, but we call that process sequence validating.
Unknown Speaker 23:37
So I'm just curious if there's certain guidelines you follow when you are validating that barcode, so say, You, you, you know, paste it in all the databases, and I'm just making this up, because I've never done it, but
Unknown Speaker 23:53
say, you get it like, is there ever a case that you get 100% match? Or is it more so like, 99? Point, whatever, then you have to make that decision? Like, are you getting 100%? Or were those percent matches? Oh, yeah, absolutely. All. So there's two other collections that people made other universities 100% matches to types? Definitely, those look good. So is there kind of a rule of thumb of, you know, I don't get 100% match, but I get, you know, what if it's 99 point, whatever it what, what classifies that as, oh, this is a subspecies. This is a new species, but same genus.
Unknown Speaker 24:37
Like is there a rule of thumb of if it's 70% match, it's the same genus, but new species if it's a 95% match, it's it same species, but a subspecies, something like that. I don't know if that question makes sense. i Yes. I understand the question. It gets a little complicated. So follow with me here. So in the DNA realm, if like you had mentioned 70%, so 70 Plus
Unknown Speaker 25:00
Seeing is way, way far away. So you would tell me
Unknown Speaker 25:04
to always so right, because like bananas and mushrooms, we share like 50% of our DNA with Yes, that's it. Yeah, yeah, it's a Yeah, that makes sense. In the particular philosophy case, the closest match was 95%. So that also is pretty far away. And usually what happens is, you can least match it to the genus. So it's like, giving you you know, obviously 100 100, you're like, Okay, that's cool. If you have like, 99.5 or higher, it's probably still 100% match. There are lots of different exceptions. And really, I think there will be a standard put out, but I don't know if that I can really speak on to that standard yet, because we're still learning so much. And, and there's, yeah, this, it goes, it goes hand in hand with the whole what is a species. So it's really like, it's obviously at the end of the day, it's humans making these decisions. So
Unknown Speaker 26:01
99.5, and it's probably still 100% Match minus a couple small things like you have, sometimes you have to enter IU PAC codes, which are basically two nucleotide acids, claiming the same position. And that happens often. And sometimes, the person that's doing sequence will enter the IU pack code, sometimes people wall, it just depends. So you got to kind of look out for those things. 97. And down, we usually call like, it's not that species, for sure. 98 There's a it's like back and forth. Like there could be some miss
Unknown Speaker 26:41
some messy stuff and errors in the sequence. And that's why we're short a little bit or just something that's needs, like different gene region sequence to better your state. But I'd say under 97, it will usually say that's a long way away from the species. But again, it's kind of hard to answer that completely without like seeing the process of doing it. Because every, um, you know, every genus is slightly different, too. So.
Unknown Speaker 27:09
So um, I'm curious, because
Unknown Speaker 27:14
like Homo sapiens, were the same genus and species, right. But our DNA probably varies quite significantly.
Unknown Speaker 27:24
How is there ever 100% match? Would it be a clone? Or would it be like a species of fungi that
Unknown Speaker 27:32
they reproduce to have, you know, they're literally clones of each other? Does that question make sense?
Unknown Speaker 27:39
Sure. We didn't get through it either. So my understanding is, like, obviously, not every gene region can we use to get to these same answers. So the IPS region is very conserved in fungi. So that one in particular, as well, and I, that one's targeted? As far as like humans, there's a different gene that's used. It's one of the Cox genes, and I'm not sure not too positive, but maybe a little more curious. I'm sure that that's kind of the same one so unsure, obviously, yes, we are the same genus and species, but all of us are different, our DNA will not match.
Unknown Speaker 28:19
So let me think on it, and if I get back to an old couch, I'll catch back up with her. Yeah, cuz I was just interviewing someone talking about how
Unknown Speaker 28:29
death caps can produce bisexual or unisexual Lee and so they can basically create clones of themselves and they only need one spore to make a new mushroom, but it's a clone of the previous but they can switch back to, you know,
Unknown Speaker 28:48
a hetero, hetero, periodic sperm carp so they can, you know, obviously, each each of its offspring is genetically different. So they can switch between being a clone or, you know, genetically different and apparently, a lot of mushrooms do this. And, you know, if you talk about yeasts, they're you know, it's the same
Unknown Speaker 29:09
there it's basically a clone.
Unknown Speaker 29:12
So yeah, I that that's it's really interesting that
Unknown Speaker 29:17
yeah, I even read an article about
Unknown Speaker 29:22
something about like a, I think it was a crocodile or an alligator that did this in captivity, where a female alligator I think it was that gave birth without any males around and it was a genetic clone to the mother.
Unknown Speaker 29:38
So yeah, I don't know I don't know if that's what's going on. If you get 100% match of that proves it's it's reproducing Yunus actually, or I don't know, maybe I'm looking into it too far. That senior answer,
Unknown Speaker 29:52
but I'm sure when this podcast gets posted, somebody will let us know for sure. Sure. With all the amount of listeners you have someone knows the answer. Yeah, yeah.
Unknown Speaker 30:02
So, when you started, you know
Unknown Speaker 30:08
I hear nanopore technology, a lot of people are raving about it. It's a semi new technology that is kind of revolutionising
Unknown Speaker 30:18
cheap, easy DNA sequencing for people that is making, you know, it used to be, I'm just making up numbers like 15 bucks a test, but now it's like cents.
Unknown Speaker 30:30
And
Unknown Speaker 30:32
so I'm just curious, like to get started, if you want to make your own DNA lab or your lab like, what, is there a starting kit of different tools that you might need? And then what are some upgrades that, you know? How much how much are we talking? How big are these machines? How easy are they to get? What tools do does one use? Both both just beginning and then if you're like, This is my life. And I don't have the top tier stuff like what what do we what are we talking about? Okay, yeah. So if you were Are we just strictly talking about nanopore? Do you want to mention Sanger sequencing? Yeah, I don't know. Yeah. First, let's start off with what is the difference between nanopore and you say Sanger tech or sequencing? Yeah. So yeah. Again, I'm not a complete expert. So I'll give you my best explanation. So Sanger sequencing is like the traditional and first way of sequencing and they use different machines. And for my understanding, like I, myself couldn't purchase some machines or two large, huge contracts. But basically, those machines, and I don't know the methodology, but they're reading now, there was at C's and G's of, you know, the
Unknown Speaker 31:50
PCR material that we send into the company. And you were close to right, like it is about $15 per sample, maybe a little less, but it definitely could be upwards to a little over 20. So that was like just for one sequence. That was, if you did that extraction and PCR whole, then you would have to send off for sequencing. So with both nanopore and Sanger, you still have to do pretty much all the same steps, you still have to extract DNA, you still have to amplify the DNA in the gene region that we're targeting, which is always in my case, it is for right now. And then you have to sequence with Sanger, you then would have to send it off to a lab. So in my case, I'm gonna send it off to California. And to do that, I think I was getting it down to about $7 and sample here. And that was with my extraction, my PCR and then mail limits California and averaging and our bell $7 per sequence.
Unknown Speaker 32:56
And to get started with that, yeah, you mean, one could mean, they would give you what would they give you back? Just the barcode? Or would they do some of that manual work to call them in like a text file, and then you would get a chromatogram and it would be a TCGA? TC Chingy. So you still have to, you know, put that into databases? And do all that work? Okay, yes, they'll just stick in my market and you'll go right into the sequence validation process, a sequence validator would then have to start taking those files and going through them for to get started on that. I mean, 1000 1200 to two SCRAN could get you started. I think like if you really budgeted in singer so yeah, so you get 1000 to 2000 and then you'd still have to pay seven bucks a sample then do all that manual work yeah, you're gonna fail price per sample. And you know, most of the stuff like only consumables, plastics, lab tools, as in like, pipettes and stuff, I mean, you can find it all on that game Amazon, you can find it on other websites online super easy. As far as like thermocycler is and stuff, you can buy them brand new, and they are very costly or what myself, and I was recommended by the emojis Alan and Steven and Hart and all these people is like EBIT. So I bought most of my thermocycler is off eBay, you just get on eBay. And that it I think I've seen one before you just put all the little tubes in like a circle, then it spins around really fast and homogenizes the word or vibrates or whatever it is that you're actually that's the center, okay. Yep. And you will need one of those.
Unknown Speaker 34:41
But it's not if you're if you're starting to actually budgeted way you actually don't need a centrifuge. But that's all I'll get into that in a second. So you know, 12 letter to a couple grand. gets you started saying your if you're budgeting and get a couple pieces of equipment off of eBay, and you can get these things off you
Unknown Speaker 35:00
because universities and labs all across the country are constantly upgrading and getting right, we're, you know, closing down or whatever. So, most of all my machines have had University stickers on them or whatever. So, yeah, I got I got all my flow hoods and back to incinerators and all that stuff from eBay, that's yeah, you can get a lot of cool stuff there. It really is. It's all the science stuffs on there.
Unknown Speaker 35:23
As far as nanopore sequencing, again, you have to extract, you have to amplify, and then you have to sequence but now I'm gonna seek not sending any samples out of my lab. But, you know, basically, before I would have these two laid out the green solution, which was the polymers and the primers, and basically, they would do all this fancy stuff, cleaning it up, and then you know, get the DNA all there and sequence. Now, all that is actually done here. So I have to clean it up myself here in the lab, I, you know, get it all the way down to a specific concentrated DNA, basically, very, very pure, concentrated DNA. And I load that into the New Oxford Nanopore sequencer, and everything's done here, house with my computer in that device takes about 24 hours to two days, 24 to 48 hours to run. And then the data is now added on to my computer. And from there, I still have to do what we've talked about the manual, grabbing blast, teen looking and making that call and updating stuff. Now we do have some really cool tools like myco map, which Steven created. And that connects with I naturalist and GenBank. So like, you can do a lot of your work and myco map. And then it pushes out the information to the other databases all very, very efficiently. And it's a lot more helpful. So. So after you extract it, do the amplification, PCR. What it's just like a little, I've seen pictures of the Oxford Nanopore, and it just looks like this a little, it's telling you the rectangular thing that you plug into your computer, what do you just like drop the the green liquid onto like a plate, and then your computer just runs a program to sequence it is basically like that. So the green will actually is gets pulled out. So basically,
Unknown Speaker 37:23
if you can imagine 960, tiny little tubes of green liquid, you condense them into one. And then with magnetic beads and some other cool processes, you basically clean that up, and then you work your way down to a very small, almost unimaginable melt. Have you ever used the pipette by chance? Yeah, yeah. Many years ago. Yeah. All right. So a suit, the smallest pipette tip we usually use is 10 microliter pipette tip. And
Unknown Speaker 37:57
in this particular case, we are getting down to seven microliters of the DNA, that sub concentrated and purified and now gets loaded in onto a flow cell.
Unknown Speaker 38:11
That's what that's part of the device. And within that Oxford Nanopore little USB thing, and if you are right, it is a very tiny, and you're loading seven microliters of up to 960 different DNA samples onto the device, and so on. So you can you can find 960 mushrooms in the wild, extract all of them do PCR amplification, and then run 960 different mushrooms at the same time. Is that right? Yes, that's correct. Yeah, obviously, remember, there's failures at infinity? Wide. So we'll get there. I'm curious, like, how can you mess it up? You know?
Unknown Speaker 38:52
But
Unknown Speaker 38:54
then, so how much is that Oxford? You know, I think I went to their website one time, and I saw they have like, different models. And, you know, some are more expensive than others. And I'm guessing the more expensive ones can just run more samples at a time. Is that right? Yeah, that's pretty much right. But it would never really been necessary for what we're doing in fungi. Those are like, for COVID and virus, then quick mutations and human genomes and stuff. So yeah, the Oxford device itself, that little itty bitty device is only $1,000. And when you think about it, in terms of anything else, scientific and throughout all these years of sequencing, it's actually almost another thing. It's unimaginable that the device itself has only $1,000 when traditional sequencing machines are hundreds of 1000s of dollars, right, right, and maintenance contracts.
Unknown Speaker 39:48
So super affordable, but again, people I don't want people to hear this and be like, Oh, it's like 14 grand to get started. But you said 1000 Well, the device is 1000 the sequencing like device
Unknown Speaker 40:00
So 1000, there's still a lot of other costs that go into basically making a complete nyrup or setup. However, I would estimate anywhere from eight, to 10 to 14k, just all depends on a few tools. Now, I do know that I did try to go the cheaper route. And it wasn't really working all that well, my success rate. And then as I invested and spent a little bit more money on a couple more machines that I needed to really hone in a few processes, my results are starting to show. So
Unknown Speaker 40:37
the device itself, yes, it's 1000, super affordable. But just everyone needs taken mine, there are other costs. But I would say you have close to 8000, you could definitely get started in that 1000 would include you to like literally run for probably 10 to 20 runs in runs, meaning like these actual sequencing rods like where I'm heading into whatever right ready plate, so there's 96 in the plate.
Unknown Speaker 41:04
And if you do,
Unknown Speaker 41:07
what is a 10 plates here at 960. So I have not personally done 10 plates yet Stephen Russell was the king of doing 10 plates. My Max is eight plates. But I'm just slowly working. It just says I continue to get more comfortable and familiar with my process. Yeah, that one? Eight to like 10, I would say most to like 14. If you think about it, it's it pays itself off in the long run. Because now you're taking that seven to, I would say seven to $20 sequence. And you're literally taking it to cents, right? Like I mean 30 forced and just the cost of that plastic tubes and the the Yeah, and we can feel it because it all like it's a little bit upfront expensive. But when you break it down, like everything goes a long way, usually. And yeah, it literally I think the cheapest sequence is 34 cents. Now, I'm probably a little higher than that, because I'm using I'm gonna follow up everything to the tee and using some cheaper stuff. So maybe I'm still within under 50 cents probably a sample. Would you get the analogy that, you know, this the Sanger technology is just archaic like is it
Unknown Speaker 42:27
does Oxford Nanopore just is it just better all around? Or is you know, I mean, like computers back in the day used to fill up an entire room, and they could just do the functions of what a calculator. And now we have, you know, the most advanced technology and are in our pocket? Is it? Is it related to that of just the technology is just better now? Or is it a cheaper alternative that say, you know, Sanger is more reliable. And that's why it's more expensive and nanopore is just your percentage that you'll fail is just a lot more, but it's cheaper is that that's really accurate. What you just said there at the end. So technically, I think most scientists mycologist anybody in this field would say Sanger sequencing is actually still more accurate. Got it. So but like you just said at the end your your throughput so high that if you're fail Ray is isn't really going to affect you as much because the throughput so high, and we really can never imagine getting like 780 sequences back saying you're at a time. But we are doing that with nanopore. So like Stephen Russell was running 960 He's the guy is doing 10 plates. I think actually, he's just now cut down to eight plates. And I can explain why later, but it's because of a robot. But before the robots even was doing 10 plates, and he could get a lot of robots are stealing our jobs, man.
Unknown Speaker 43:59
They aren't stealing our plates.
Unknown Speaker 44:03
Kids are really kind of agree with it. Because of yeah, just the amount of pipetting that one does have to do in order to basically yeah, that 160 is insane. Yeah, it's a lot by 30. So how can one mess up? Obviously, his DNA, so I'm guessing your own DNA can contaminate or other other DNA things can contaminate your samples, right? Yes and no.
Unknown Speaker 44:30
So there's without getting too into it and making it complicated for the audience. There's a couple different ways that we utilize primers when sequencing and it's different between nanopore and Sanger. And so, contamination is traditionally a big risk and sequencing specially Sanger sequencing because we're just grabbing an IGS standard primer and trying to
Unknown Speaker 44:59
base
Unknown Speaker 45:00
SQL, you amplify that by ETS reading from the fungi. So anything else that contains ATS that gets contaminated, you know what I mean? It could like pull that in and cause issues with the nanopore sequencing, they use tagged primer. So, say another organism gets into like that well, and what actually is a good example is like, like an AMA and EDA, that's,
Unknown Speaker 45:25
you know, being parasitized. And so the,
Unknown Speaker 45:29
if you take a small super small piece of that specimen, and you put it in the to, with our nanopore sequencing primers that are tagged,
Unknown Speaker 45:37
if everything goes smoothly, which usually does to sequences will spit out the machine and onto the computer and files. Put in that position of that well, so then you're like, Wait, I got it two sequences? Well, it's because you're getting the, the host and the contam off of the targeted sequence. So contaminations, not that big of an issue compared to Sanger. But I definitely would say it's still an issue, I've definitely noticed it. So you can mess up by that. You can mess up by the simple common human errors of like, fat fingering stuff, like typos, like putting the tissue in the wrong to like messing up your orientation. Now, it's all like beginner stage stuff that you can mess up. But you could mess up almost everything,
Unknown Speaker 46:29
just because it's following so many steps. So it's kind of like baking a cake with like, 200 ingredients. And yes, it feels all lined out, you can follow it. But like, if you're just get a little bit complacent, or you're a little bit distracted, and you forget, like one ingredient. It's basically over. And that's kind of how it is with snail poor sequencing, especially when you get into the back end of like the ligation and adapters and stuff, you're just basically adding stuff together and following volumes and following incubation times and stuff like that. So you could mess up there.
Unknown Speaker 47:05
But that doesn't usually happen. It's usually at the very beginning, you're gonna mess something up in the extraction phase somewhere.
Unknown Speaker 47:13
And then as far as like sequencing and loading the flow cell, it's pretty easy just to watch out for an air bubble when you're doing the like the actual loading with the pipette tip. But that seemed kind of stressful in the beginning, but turned out to be super easy. So yeah, there's it's just really when you get those pipettes and the tools in your hands, and then when you're typing numbers into a spreadsheet.
Unknown Speaker 47:36
So I'm just I'm just kind of running some numbers you said, you know, startup costs for singers have around, you know, two to 4k to say average street 3k.
Unknown Speaker 47:50
nanopore is
Unknown Speaker 47:53
between eight and 15. So, average like 11 and a half K.
Unknown Speaker 47:59
The difference between those two, like on average is like, like, eight 8500. This is like, I'm just like writing some quick back of the napkin things. It's about $7 per Sanger test to send it in.
Unknown Speaker 48:15
Why would say like, you should probably go to Sanger unless you're running. Yes, like 1200 test. If you were running over 1200, you should probably go to the end and pour, because over that amount of tests would probably be cheaper. So if you're just starting out, just kind of if you if you're not doing 1000s of tests, that maybe maybe stick was saying GRE and and that is the cheaper route. But if you're doing over 1200 tests, approximately give or take, you know,
Unknown Speaker 48:48
then invest in nanopore long term for sure. Yeah, if you want to learn sequencing, and you're just super excited to sequence your stuff and your friend stuff, and you're like, Dude, I have not going in like Kyle and Steven in heart, then you have to go to singer because it really is fun. And
Unknown Speaker 49:06
it always gives you that set up the way if you do want to go nanopores, you really understand it a lot more in depth.
Unknown Speaker 49:13
But yes, if you are planning on sequencing a lot of mushrooms, you should go the other way. So just to give you I just did some quick numbers. On my naturalist
Unknown Speaker 49:24
project, we have 15 144 Good sequences that have came through. Obviously, we've added some that failed. So I've read over 1544 in it. Yeah, you're in the green, you're in the Yeah, if you times that by sudden, you're 10,800. So
Unknown Speaker 49:42
the cost to do those 1500 is almost the cost of the nanopore lab. So you look at it in context like that. It's like, well, this, this really is it's just the upfront cost. That might discourage folks. But again, I wouldn't really recommend going this route.
Unknown Speaker 50:00
unless you have huge plans to mass sequence stuff, or you're going to get into other sort of, like sequencing stuff, or maybe you're interested in full genome, that'd be a good way. But if you are like, Oh, I'm just going to sequence like, couple 100 things a year, then I would definitely either send it into someone or just go singer on your own, I definitely wouldn't want to invest, you know, upwards to 10 grand to only do two other mushrooms a year or something, you definitely want to do 10,000 a year if you haven't no fun. So, Kyle, you're offering this for free?
Unknown Speaker 50:37
How are you doing this? Why? Yeah, how am I? Good question. So, man, I'm just a guy who likes freedom. And I feel like if, if if the money is out there, and it can be utilized for other things, why not? And, you know, I after the philosophy thing I was super interested in Allen did it for me for free? Alan's absolutely the best. He did it for me for free anyway. And that kind of just inspired me. And I know that obviously, Alex can do it for free from the whole world. And, you know, not can many people but that is kind of the goal that we are trying to say is like, see, if you want something sequence on there, there is there is a route. And so yeah, I've used some of my own funds quite a bit of a really to kind of get started. And then yeah, I just started doing crowds, crowdfunding,
Unknown Speaker 51:33
that people were pretty responsive to it. And, and the results kind of started speaking for themselves. And honestly, yeah, I think the the donors and the people that are really into this, they're, they have no problem throwing some bucks. My way, knowing basically what it takes, and then seeing the results kind of unfold. And yeah, I use 100% of the funds that I gained from friends and family and it goes straight back into the lab.
Unknown Speaker 52:02
My lab is completely equipped, I'm fully invested, I just still have costs like reagents reusable, and stuff like that. So you still gotta buy flow cells, the flow cells are not reusable. And then obviously, all the chemicals in the the consumable plastics and tips, the tubes and stuff like that they are free is where I really want to stick. I want to make it free to the public as much as I can.
Unknown Speaker 52:29
Just to kind of set the standard. So that's what we're doing towards, have you heard of the, I don't know how new it is. But it's kind of like a
Unknown Speaker 52:41
docu series, I wouldn't even call it a documentary. It's kind of like,
Unknown Speaker 52:46
I don't know how you would classify it, but it's about the start of Spotify on Netflix. No, it's good. It's, but like, I didn't know how it started. But all the founders, apparently, they wanted to make streaming music free. And this was like around the time of pirate bay and Limewire where they were doing it illegally. And so they were the first streaming company that made music free, but legal. And, you know, they had a they had to incorporate, like, Spotify Premium to have a business model, obviously. But like, their whole, you know, their founders were like, very much like, we need to have this free or, you know, like that. That's our ethos. And that's what we need to do, like,
Unknown Speaker 53:34
but it's good. I said, I started watching, I was like, wow, this is pretty cool. I didn't know about that story, but it kind of reminded me of the way you're talking about this. So I really appreciate that. And
Unknown Speaker 53:45
yeah, that's, that's amazing that that you guys can offer this this service for free and, and support the community to continue understanding, you know, what it what is going on with fungi and mushrooms and
Unknown Speaker 53:59
and find new species like that philosophy and, and many others. And, you know,
Unknown Speaker 54:06
obviously, I've heard many different ratios, but I think it hovers around, we've described, like 1% of all fungi in the world. So the more DNA sequencing we do, the better.
Unknown Speaker 54:21
And yeah, and obviously fungi have a lot of different benefits for solving a lot of our our biggest world problems. So if we can find new species to do,
Unknown Speaker 54:35
not only DNA sequencing, but But you know, figure out what compounds are in them what what uses that we can use for like myco, textiles or farm, new pharmaceuticals or new whatever. Next, that's the next thing I almost guarantee that that's what's kind of what we're setting up and leading towards you.
Unknown Speaker 54:55
So what what would you say is the hardest part of this work?
Unknown Speaker 55:00
So the hardest part is tissue collection.
Unknown Speaker 55:04
And I say that because it surely is the hardest part, because it's just so time consuming. It's very tedious. And there are many
Unknown Speaker 55:16
potential errors in Bolton tissue collection. So that's one of the harder parts and then just maybe yeah, keeping up the
Unknown Speaker 55:24
keeping up with the public and keeping people happy with getting results in a timely manner.
Unknown Speaker 55:31
And the flip side, what, what keeps you going and offering this the services and despite the, the high upfront cost, and all the time that goes into it, and the reoccurring costs and everything? Like what what keeps you passionate to keep wanting to do it and offer it for free and help the community? Yeah, good question. Yeah. So first and foremost, my love and passion for my college is just out out there really, really far. So, again, with old philosophies of moraine incorporated into my college now like, okay, how can I make this accessible? And when we did, and I say we, because I have a great, great, great volunteer team
Unknown Speaker 56:17
that also agree and kind of like the way that we run things. And they actually work on the team and do quite a bit of work. And they do it all for zero cost. And they know that, you know, there there, there is no funds for me to be able to pay people to do this work. And I, at the same time, I don't want to seem like I'm exploiting, because that's not what we're doing. Like, we have our own little group. We work when we work, and we just try to get stuff done. And every one of these people that still on the team, are super passionate people about mycology. And yeah, they're the same. They're kind of the same as me. They just wake up and they're like, how do we make this known? How do we work on this together? And how can someone straight off the streets, Allah or the forest out of anywhere, participate and fall in love with this? And that's really the way that we approach it. And that's to me, it's super motivating. I wake up every day and I'm like, mushrooms, Dean agrilus. Go like that. After this podcast. I'm getting busy. I promise the team OMD. Oh, nine, so that'd be my ninth one. So I am working on eight plates for that. So there'll be super excited to get, you know, 700 or so mushroom results back so they can work on? Hell yeah. Yeah, I tell people all the time, you know, if I could clone people like Elon Rockefeller, and now you I would I yeah, I would amplify you guys, you know, copy and paste that DNA all day. But yeah. Yeah, we, it's, it's exciting it. I meet people all the time that come into the mushroom space, and they go to a mushroom festival or whatever. And, and they're like, what I, everyone's so inviting. And everyone's so helpful for each other. Granted, there's some, you know, some always some bad apples, but, but the majority of people are just really friendly. And they want to help each other and they want. They're excited to get to teach people about this and excited to share information, generally. And the noon. Yeah, no, and it's just,
Unknown Speaker 58:27
for me, it's been refreshing. And I love interviewing, you know, people like you and so many others. But it's also refreshing to hear outside people from different industries, you know, get introduced to the mushroom space. And like, really, like people are unbelievably cutthroat and competitive and really, like, closed off. And, you know, and there is some of that happening. But for the most part, people are really great. And they just want to like, further the passion of mushrooms and mycology, it's just, it's just so it gives me faith back in humanity again, of of, of what we are capable of when we have a combined passion for something bigger than ourselves. So good job. Keep it up a lot of you out there. Yeah, we will. We'll definitely keep it up. I don't know if I mentioned Alan is on my team. So I'm honored to have him on there. Alan works as a sequence validator and pretty much an all around boss that just helps answer your questions and stuff.
Unknown Speaker 59:30
But yes, you're right. Majority of the community is like that, like, I mean, countless zoom calls, phone conversations, text messages, Facebook emails with some of these top people and you Yeah, just nothing but the utmost respect. And they answered all my questions ultimately helped me and they honestly, I'm a paid full tech person. So I'm now in a position where it's on pay forward. I want to pay it forward. If someone says, Hey, I have all the funds. I want to start this right now.
Unknown Speaker 1:00:00
I have no problem sitting with them as much as I can and calls and saying, Hey, that's what you got to do. This is what I did whatever works. Um, because ultimately, that's just the way it should be. It's just pass on the knowledge, as much as you can continue generations and other people working. That's how I got here. So. So if you had unlimited funds, the best equipment in the world, the unlimited team, unlimited time, you know, what, what would you do? Well, we would sequence everything that
Unknown Speaker 1:00:31
exist on Earth, and we would do it just, I don't know, in waves, or we do all the fungi first, then go to something else.
Unknown Speaker 1:00:41
Yeah, that's what we would do. Because it's like, we already have Allah own of the sequence mushrooms between the three different companies that are doing it. But we saw a lot lot more to go. But it's safe, whether we have funds and, and resources were not there. And maybe we will be able to knock things out quicker and move on to other cryptic groups like insects and other things that need sequencing.
Unknown Speaker 1:01:06
So I'm curious if there's like a style of, or like a category of, of fungi that you feel like
Unknown Speaker 1:01:16
you're most excited to sequence or you feel like there's not enough information like you brought up, you brought up insects, I'm a big fan of entomopathogenic. fungi or fungi that attack insects are core to sepsis. Most people talk about them, or, you know, like we brought on someone talking about marine fungi A while ago, and I haven't really found anyone since that is into it. And it seems like that's another area we've discovered what 1% of the ocean and then marine fungi. It's another like, we don't know anything about that. But it's 70% of the the earth is water, you know, your or like, I don't know,
Unknown Speaker 1:01:53
endophytic fungi or like, there's a bunch of weird fungi that exists. Like, I don't know, if you're super excited. If there's one category that you're like, oh my god, we would love to sequence that all day. We'll read those, isn't it? Maybe biased because it's awesome happens to be like my favorite genius mushroom but it'd be at the Naseby genius.
Unknown Speaker 1:02:16
Yeah, maybe mainly because it's my favorite. But also it's just because
Unknown Speaker 1:02:20
they are so cryptic. And I'll
Unknown Speaker 1:02:25
cap cat we're up to so many
Unknown Speaker 1:02:28
nonprofits have an awesome I mean, to me, it's just like, kind of like a no brainer or want to attack like,
Unknown Speaker 1:02:35
what's got the biggest workload first. And that's also interesting, because there are things have bigger workloads that are just not as much interesting, at least to me. So anosmia actually worked for I got on this call, which was looking at an ostomy that I found an old growth forest. And Alan was my sequence validator on that one particularly, and he gave it a new nonprofit on Ohio three, so we give these shortcodes with the state and a number instead of like souI arises Soulier Ryza is a nonprofit that's had a little bit more time and thought to it. The are just working on the fly and we're not really spending too much time with these things. I'll tell you the three
Unknown Speaker 1:03:15
so makes it easy. Yeah, wants to come in and put a good Old Navy one at all. So nice to be and then my second may be m&e Does m&e is really really cool. A lot of cool stuff coming from them as well. Sweet. Well, where where can people follow your work? Where can people support you?
Unknown Speaker 1:03:33
How can people send in samples if they want?
Unknown Speaker 1:03:37
Yeah, give us the loadout Yeah, cool. So my biggest thing is Facebook comm on Facebook, anyone can reach out to me anytime I do tell people that I have kind of like a high influx in my inbox of volume of people asking questions, but I have links for everything like in my Facebook. So if you go to my Facebook, you go to my all links should direct you straight to a protocol, that protocol if you like to three page protocol, there's some really cool pictures in there. I just recommend anybody that wants to get into it. Like if you want to send me specimens to sequence, I ask that you read the protocol at the bottom of the protocols when you address and you get shipped according to the protocol. And then just give me patience. I do probably have upwards to 5000 mushrooms sitting in my in my office now that are that are in queue. So quite a bit of mushrooms in queue.
Unknown Speaker 1:04:27
I do plan on getting into every single one of them. And so that's how people can send them in. Yeah, just Facebook and I have a Facebook page that they could follow to which is the Ohio mushroom DNA lab.
Unknown Speaker 1:04:39
And if you slip in like a five or a tenner in there, you might get it to the top of the pile a little quicker. It's pretty
Unknown Speaker 1:04:48
trucks and stuff. I'm like, I gotta get this done for them. Cuz you know, ultimately money does talk to and I don't want anyone to think that their donation wasn't greatly appreciated because
Unknown Speaker 1:05:00
They all are. But I also do try to keep it kind of fair to the people that are sending money.
Unknown Speaker 1:05:06
Maybe we've had a discussion before, but like, yeah, or whatever. But, um, yeah, it's just it will take a little time. But we are working pretty hard. And we are working towards
Unknown Speaker 1:05:18
finding some lab hope and going towards just as many runs we could possibly do in a year. Sweet. Well, thank you for for coming on. And it's it's great to hear people like you doing this work and expanding the field so we can understand just a little bit more of what's going on and discovering all these new species that help us greater understand mushrooms in a in a bigger way. So Thanks, Ben. Appreciate it. Thanks. Thanks for having me. Great. Thanks to, you know, the podcast and podcasts in general. Hopefully, the information gets out there. All's we're trying to do is just spread the love. Put our two cents in where we can with mycology and just hopefully we can help someone in the future.
Unknown Speaker 1:06:03
Cool and and thank you everyone for tuning in and tuning in to another episode of the mushroom revival podcast. We hope that was an exciting episode. And if you learned something today, yeah, tell tell, tell a friend to a family member. Maybe hopefully this inspires you to do some DNA sequencing or send in some samples. Get out there maybe find a new new species. And you know, if you if you want to support the show, we don't have a Patreon are any way that you could donate directly but we do have a brand mushroom revival that has a bunch of organic functional mushroom supplements from you know gummies to capsules, to powders to tinctures. And so if you if you want to get something for yourself, we have a VIP coupon code just for listeners. And that coupon code is pod treat for a surprise discount code we change it all the time. So you got to figure out what you're gonna get. If you want to spend any money we have a we have a giveaway going on the link is in the bio so you can when we pick a new winner once a month, and you can pick whatever goodies from from our site. We also have a bunch of free educational content on there. We have a ton of blogs, we have a bunch of free ebooks, but your recipes and you know things about psilocybin things about fungal ecology, you name it, everything mushrooms, and also my newest book, The Little Book of mushrooms is is on there as well as well as pretty much anywhere you could you could buy books like Barnes and Noble and we're in urban outfitter books, book section and a bunch of others small book shops and stuff like that.
Unknown Speaker 1:07:42
Or you just get it from from our site. And apart from that, you know, thanks for tuning in. This is a beautiful mushroom community and sending sending all the blessings out and as always much love and may the Force be with you
Transcribed by https://otter.ai
