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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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Mycelium develops substrate specific digestive enzymes in order to digest what it is on, If you culture only on MEA, eventually just like us the myc gets "bored" with the same old same. If I know I am doing a specific grain master the agar for the mother plates on the spawn run gets a pinch of every thing that the myc will encounter in that planned expansion. If the final sub is wood, I obtain a sample of the intended wood and a pinch goes in the pot. This may be over kill in some viewpoint, But I have demonstrated to myself and it is also reflected in the literature that "leap off" time, or the time between inoculation and the time aggressive capture starts can be delayed by the myc having to stop and develop digestive enzymes for a sub that it is unfamiliar.
keeping the base grain and the source of protein changing challenges the myc and encourages archetypal growth. To me this is why LC and specifically fermentation is so important in a big production run, also in other instance, a questionable sterile arena, this enzyme preparedness may be the difference between a more rapid colonization that overwhelms minor contams preventing failure, by not allowing 24 to 72 hr for the contam to get a leg up. Nature is all about niche and dominance, when we create a sterile vacuum for a myc to colonize it sometimes is that a perfect job is not done, in small percentage I have seen a tiny spot of trich or asperilligus get overwhelmed and killed by a rapidly colonizing myc. In reference to your aspirations of bulk outdoor substate left untreated, this could make a difference that would matter. I keep a grain storage facility for makeup in a special bin, some of my favorite grains for strain work are, Rice white millet red millet quinowa wheat My favorite protein sources, High quality dog food spirulina peptone oats soybean meal Salmon/Sardenes (believe it or not) in every solution I add an extra light Malt Extract as a basic source of easy calories, The ratio of the other grains is never meant to be specifically nutritive, only to flavor the agar so the myc is ready when I open the gate for a rapid expansion. When I am mixing for slants I go with a strictly MEA with a double agar content, In that instance the medium is to no other end than grow out and storage. The double agar ensures a firm sub to work on in tight quarters and that it will stay conformed to the container. I also activate a specific wine making yeast manufactured by Lalvin 71b 1122. I use this strain becuase it nutritional requirements are low and it is easily activated on ME The culture is inactive saccharomyces cerevisiae, I add a generous amount of yeast, let to activate on the surface before agitating for 15 min or more on a stir table. This activates the yeast and then the autoclave kills them. I believe that higher fungi benefit from the fact the the yeast mineralize nutrition from the sub then making it more available to the myc. I also believe that the myc will digest said yeast both living and dead on a sub. And as always (I seem to say this allot) refer to chapter 12 of Stametes Growing gourmet and medicinal mushrooms, GGMM for further reference And don't forget the hotly debated peroxide- it makes an unforgiving substrate a little more doable for the common man. See Rush Wayne or Kerry Orgame for formula. Fun fun fun with agar Blessings and light Mycoelf -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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Coolaid smile Registered: 10/02/09 Posts: 897 Loc: SW WA Last seen: 11 years, 3 months |
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Its taken me a while to digest what you wrote. I now understand that I don't know shit yet and have a lot of learning ahead of me.
I think I understand some of what you wrote. Fermentation? Is something I don't understand. I will also refer to chapter 12. Thanks again for the schooling. -myc
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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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knowing that you don't know is the point of the wheel where knowledge becomes wisdom, and is the first step onto a new path. The rewards of knowing where this one leads are infinite.
I am a Stametesian, I don't worship the man, but subscribe to his school of thought regarding mushrooms and their cultivation. He is an excellent teacher in his many books and does not tell you what to think but merely suggest which questions to ask. To me this is the core of the scientific process. The process of mycelial fermentation is one of a myriad of forms that Liquid culture can take. The man who tutored me in mycology taught me a process called myc fragmentation. In short you take a kick ass culture in petri, transfer it as a whole into a sterile blender like device and chop it to bits using a very sharp high speed blade. There are special blenders that are sold for this that are fully autoclavable and very expensive. I am a retired metalworker and manufactured mine. The result of this is a liquid solution that contains huge quanitys of slightly damaged hyphae in solution. This solution can be applied directly to a grain substrate for further colonization. The advantage there is that one 100mm plate fragmented into 600 ml or so can inoculate 60 or so 1 quart grain masters each at a substrate weight of @ 200 g. Each of these grain masters could the innoculte from 1 to 10 5lb bags of end substrate or to create second generation grainmasters, wich then could inoculate a bulk substrate at @ 20% of volume. The point of this really is to take one really kick ass culture in run out to quite potentially hundreds or thousands of pounds of spawn. Just depends on how you design the spawn run which moves you make. To generate equivalent spawn using traditional technique 2 wedges become 1 master, then in turn becomes 1 to 10 bags. grow out is slower as the sub is not as uniformly inoculated, more leap off points, quicker colonization. you also have to generate 1 plate for every six grain masters. LC gives a tenfold advantage, and also allowing the cultivator to create large spawn runs using only the cultures in most premium form. When I cultivate a poly pore I use the traditional technique. My Reishi strains grow so aggressively a master inoculated w/ 2 wedges grows out in 4 days with one shaking, overgrows in less than 7. No real need to ramp up spawn there, as the old way suffices. But take a stropharia like Stropheria Rugusoannulata, the red wine stropheria, (also a close relative of PC) grows very, almost painstakingly slow on agar the strain that I have takes 2 to 3 weeks to make a plate, consequently the cultivator has to make many many plates to achieve enough agar to make 60 GMs, but with liquid spawn, one plate could make 60 GMs' which could fold out according to the above scenario. using traditional technique, a GM1 might take 3 or 4 weeks to grow out. Even longer in a 5 lb bag. But fragmentation is only half of the story, you will remember that I said that the fragmentation process slightly damages the hyphae. Fermentation is the process of adding your above described LC into a sterile vessel that is being agitated by a stir plate. The traditional vessel is a Erlenmeyer Flask, I have done it in a ball jar. The recipe for the media is allot like that for agar, except there is no agar, obviously because you don't want it to solidify. After a 24 to 72 hours on the table the myc has a chance to heal severed ends. every where it has healed a bud forms in the segment previous to the cut. each bud is a branch that gives rise to aggressively running myc that conquers territory after innoc so fast that thermo-genisis becomes an issue during incubation. If cultures are stored too closely the thermal insulative property of wood can actually retain so much heat that the core temp an exced 109 deg F killing the spawn. Fermentation also gives the cultivator to add a representative sample of all the substrate along the path of the planned Myc expansion. In the time of fermentation the myc has a chance to not only heal from the frag, but also to develop digestive enzymes for a specific sub that will be encountered, thereby reducing the leap off time to near zero. The remaining nutrition inside the fermented culture serves a a nutritive base for the myc to consume during the process of leap off onto the grain. I use whole grain and it takes some effort to penetrate that PC'd hull and start to gain a foothold. I am sure that there are forum members out there that frown on the process. There are really only two draw backs, the first is the danger of thermo genisis, the solution there is proper placement of cultures in a properly regulated environment. The other drawback is what intimidates most cultivators. This is an all or nothing technique!!! If you fuck up the setup for the fermentation by allowing a contam into the bottle, it multiples with even a more exponential force than using the traditional technique. when I was pioneering this for myself I once accidentally included a scarid fly into the broth. the result was the most uniformly contaminated grain masters I had ever generated. Biggest mess that I ever had to remediate also. It took a week of cleaning before I was at ground zero. But you shouldn't let that scare you, for this one fuck up, I have pulled off so many successful operations it is beyond count in my measure. This pic is Two cultures sitting side by side, They are of the same strain of a Strophariaicae. The bottle on the left is 72 hours ahead of the other, which was freshly inoculated in the pic. note the dramatic color change This is due to a massive folding out of the myc while in the bottle, due to the fact that it is in a perfect short term environment. This pic shows a prototype of the breathing apparatus that I have developed to supply sterile gas exchange to the culture. I found that passive Gas exchange was insufficient for the needs of the culture. I went thru allot of trial and error to perfect this technique as there are allot of delivery and transfer obstructions to pull off in sterile arena. I worked R+D on this for about two years before finally settling into a routine that works as good as I know how to make it. Blessings and Light find you Mycoelf -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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Bans for Pleasure Registered: 03/26/03 Posts: 42,214 Loc: Seattle Last seen: 1 year, 2 months |
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Quote: Prove it besides reading about it in GGMM. I also took his seminars, including the masters seminar ten years or so ago when he still did that one. However, I've seen no evidence of this 'memory' or 'boredom'. I use MEA exclusively and have for three decades. I've kept master slants on MEA for many years. Most of my strains are ten or more years old and have been on MEA the entire time and they still perform as well as the day they were made. I've also not seen one whit of evidence that adding some of the final substrate to the agar makes the slightest difference. The mycelium jumps off the agar to the grains and jumps from the grains to the sawdust or compost just as fast as ever. RR -------------------- Download Let's Grow Mushrooms semper in excretia sumus solim profundum variat "I've never had a failed experiment. I've only discovered 10,000 methods which do not work." Thomas Edison
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Coolaid smile Registered: 10/02/09 Posts: 897 Loc: SW WA Last seen: 11 years, 3 months |
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I hadn't thought of liquifying a full agar dish. Mycoelf you said that you will take a 100mm dish and turn it into a 600ml LC. In order to do this do you have to place the liquified agar to the flask on the stir plate? Also do you add sterile water to dilute the solution so it isn't so thick?
I thought that I had read that when making agar you want it less nutritous than the spawn so when it is introduced it will grow faster because of the nutritional boost. I didn't know that mycelium has to create a new enzyme each time it encounters a new substrate. It sounds like RR has never encounter evidence of mycelium memory. I will be honest this is the first time I have heard about it. It sounds like a neat idea and plausable if in fact that mycelium has to develop unique digestion enzymes depending on the food source. Mycoelf have you noticed a signifigant difference it leap time? I have been growing reishi and grey oysters on cardboard and I have learned that not all cardboard is not the same. Nike shoe boxes work well, the cup holders from fast food restraunts easily attract mycelium. Cerial boxes don't seem to work very well though. I was thinking if I was to soak some of the cardboard in water and then use the water to make the agar or maybe soak the grain in the cardboard water with some gypsum and coffee. What do you think?
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Bans for Pleasure Registered: 03/26/03 Posts: 42,214 Loc: Seattle Last seen: 1 year, 2 months |
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Quote: Actually, it's the opposite. Mycelium grows fastest on a less nutritious substrate. That's why a 40 pound straw log can colonize faster than a 3 pound manure substrate, even though the manure is inoculated with a spawn rate of 1:3 and the straw inoculated at 1:20. Mycelium doesn't have to evolve to grow on various substrates. That seems to be one of paul's pet theories, but I've never seen it in action. With shoe and cereal boxes, etc., that's chipboard, not cardboard. It's the final recycling process and can't be recycled again. I roll them up into logs and use them for heat in my wood stove. They burn almost as long as wood logs. Most of them will support mushrooms though, especially oysters which aren't picky at all about substrate. RR -------------------- Download Let's Grow Mushrooms semper in excretia sumus solim profundum variat "I've never had a failed experiment. I've only discovered 10,000 methods which do not work." Thomas Edison
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Registered: 06/14/06 Posts: 4,039 |
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To be fair to Paul his theory is not that far fetched, its similar to operons.
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Bans for Pleasure Registered: 03/26/03 Posts: 42,214 Loc: Seattle Last seen: 1 year, 2 months |
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Paul's a long-time friend. I wasn't attacking him, just disagree with that theory based on experience.
RR -------------------- Download Let's Grow Mushrooms semper in excretia sumus solim profundum variat "I've never had a failed experiment. I've only discovered 10,000 methods which do not work." Thomas Edison
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Registered: 06/14/06 Posts: 4,039 |
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I know you werent trying to buckle his self esteem. I havent noticed any difference either by adding sawdust or horse shit. But if this theory is in fact true then I wouldnt be that surprised. I can see why Paul thinks this.
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Coolaid smile Registered: 10/02/09 Posts: 897 Loc: SW WA Last seen: 11 years, 3 months |
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When using the chip board for oysters do you roll it up like a log or just tear it up? The first few bags Ive done have smelled really bad after a few days is this normal?
I noticed that the mycelium both from Ps. Cyan has begun to grow over each other kin a layering form on the agar. I had hypothesized that they would meet being from the same print and just merge creating dikaryon. How off am I? Are there spores from the same print that are not compatible for mating? -myc
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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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Quote: I can't say that I am prepared to prove my observations in an empirical sense, I would have to set up some serious control, work over hundreds perhaps thousands of generations of transfers, develop strict protocol for appearances, then go and analyze which enzymes were in every sample. I wish I could prove it, if i was a giant corp or the gov, Id' would just cut a huge check and wait for the report.. Respectfully I can say that I can only attest to my own experiential observation. The man who was my first teacher was basically a hobbyist who worked quite prolifically with one particular strain of PC, a Golden teacher. He was working with a hood on agar and was fragmenting cultures and inoculating PF style jars with the slurry. His sense of genetic management was a continuous descent of transfers onto the same store bought agar. He made no note of the P value system, but made a new generation every couple of weeks. He said that he noticed no problems or changes with the myc over that period of time. His technique was to specifically innoc each dish with as small a fragment as possible. The myc responded from this position with four perfect fans on quarters of direction, corresponding to the four sides of his agar cut. his solution for the inevitable genetic crash was to refer back to a spore print. That worked for him, but was not satisfying enough to me, so I hit the books and the main bible of study was of course, GGMM. Well, the prover proves what the thinker thinks. Having taken late gen cultures from the aforementioned strain I was convinced that the behavior that I was noticing (a degradation in to cottony sectors) was a result of poor nutrition. I was incorrect, but as the part of me that thought this wanted approval from the part of my mind that validates, I issued a vain attempt to "save" the strain. I started manipulating agar in make up searching for the one magic ingredient that would make my sacred strain live again. In parallel transfers from the same mother cultures, splitting the best fans on edges into multiple plates, I observed that where I loaded protein (quinowa) and High quality DFA (sardine based) the myc jumped and for a few transfers tried to resume archetypal growth. In essence it would be my experiential conclusion that diverse and challenging substrate stimulates the myc to try harder even with limited genetic resource. Point is that the same old IS the same old. What is true in the lab may not be so in the field, and struggle in the context of natural conditions ALWAYS involved diverse multi part nutrition, some of which may have been encountered previously further up the trail. It is Myc Vs. Myc out there and waiting around to find the exact combination to a nutritional lock is precious time lost. When you go out into the field looking for cool mushies to clone you are always looking for diverse intermediary habitats- places where flood, land slide, fire, cattle, whatever has created small scale disaster. Those environs are loaded with diverse origin of debris, The myc runs between points of best nutrition looking to capture the most squares first- competing in niche' with other higher fungus, tho that gets the most wins and there are acrobatics in between, gaps to jump, areas of inorganic material etc. Nature is rarely homogeneous. Also noting the mycellium is very old, as old as the first life to come up onto land, a true survivor. In that time in it's descent from pre history I am going to say that there were times in earth history that were more than challenging. I would argue that adversity is a positive pressure towards retaining the advantage of the retention of a nutritional memory. I do think that this memory fades past a factor of myc expansion. You may say with an empirical club that I can't "prove" such things , but to me all of those factors given interplay seem reasonable to allow the jump to infer that mycellium has developed a few tricks up its Sleeve. NOT having a memory for specific sub that was encountered within a perimeter of expansion would seem to me to be a distinct disadvantage, we know that Myc can run sub specific maze (i.e.mycellium running) Why does this seem so unreasonable??? Besides the observations above my INTUITION is ok with the idea of that being true. Alternative mycology, which this I presume this forum is, has got to leave room for those mechanisms that serve us where the "as taught" scientific method fails us. I am fascinated by sterile culture and have spent hours looking and meditating on specific strains. They are like little pets, an entity that is more than just an embodiment of running mycellium. They have a awareness that extend outside the petri- connected on the cosmic level. I feel as if they communicate and teach in a very silent and passive way, the cultivator who looks at a culture for ten seconds before conducting transfers and running off mentally to all else that must be accomplished in his day misses the connection- that hidden message that is there to guide us. A principal that I find valuable to be sensitive to and remains part of the myco world view that I pass on to others. All Ideas that I submit in this forum are for the stimulation and entertainment of you all. None of my views represent the be all or end all of all discussion. When I leave the established track of what the empirical book tells us it is not in spite of the established institution, but only to honor and seek out the chapters of those books that are still blank as they are unwritten. All I ask of those of you, my esteemed fellows, is that the more skeptical of you agree that a belief and the suspension of dis-belief are two separate modes of looking a singular object. You don't have to believe anything because I do, I am not asking you to by perpetuating my feelings on any given subject, but, Staying open to the suggestions of other may lead you down paths of investigation that may lead somewhere we would all like to go... The blessings of the mycellium find you, Mycoelf Ps I am enclosing a pic OF a PC Super KOH Sammui that I manipulated by placing a sterile Phalaris Arundinecia grass seed, A grass that in Nature is known to have a symbiotic relationship with some Psilo Spec. Not only did I get an amazing archetypal myc pattern, but also multiple fruiting, It is important to note that this is a 60mm plate and the mother mushroom in the middle is almost 2" long, note the spore print on upper lid!!!! -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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Quote: I have tec that is still in rough draft that I wanted to publish on shroomery, there are pics, but for now I will offer a brief over view. The flask with the nutrient solution are autoclaved for 45 min at exactly 15 psi. I made a container that is a stainless steel cup that has a tight fitting lid whose center accepts a modified cutting blade I adapted from a "smoothie" blender, the type of hand held blender made to liquefy ingredients for the drink, I replaced the bearings and modified the blade. The cup is pc'd with 150 ml of water. after cooling to below 90 F I open a 100mm dish, cut into sections, both concentrically and radially, and use the scalpel to transfer the sections into the cup. 3 one second burst followed by 1 one second burst at high speed grinds the culture into little bits and liberates thousands of hyphae into solution. The medium is then transferred into the flask via sterile Pyrex funnel, the breathing apparatus is para filmed on, then 72 hours later is delivered to whatever the next gen substrate is.. I peroxidate the culture before inoculation, and transfer via syringe, free pour, peristaltic pump etc. In answer to your second question regarding leap off, yes I have noticed a dramatic difference. Same strain, sister dish, fermentation vs fragmentation alone time difference for PC captureing a PF style jar goes from 23 to capture, over 30 to fruit, to capture in 14, fruit in 17 days. I am not a math guy but just by looks somewhere around a 50% improvement. Peace Mycoelf -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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Coolaid smile Registered: 10/02/09 Posts: 897 Loc: SW WA Last seen: 11 years, 3 months |
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The petri dish looks cool. You added a grain of rice to the dish and a couple mushrooms grew? Did you expose to fruiting conditions or just let it sit in closet? myc
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Bans for Pleasure Registered: 03/26/03 Posts: 42,214 Loc: Seattle Last seen: 1 year, 2 months |
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Quote: With the exception of peroxide, that sounds like exactly the method Paul teaches at his seminars. I take it you attended one? RR -------------------- Download Let's Grow Mushrooms semper in excretia sumus solim profundum variat "I've never had a failed experiment. I've only discovered 10,000 methods which do not work." Thomas Edison
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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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No, But thanks for the complement, it is on my wish list to do so. I know that I would benefit with a more experienced person looking over my shoulder, critiquing hand motions and technique. I would also love to qualify for the advanced class so that I could hear more theory-
I do one thing differently than Paul does, when I first tried this fermentation technique, I misread the information and added LIVE yeast, as opposed to yeast extract. I know that sounds crazy, and I am not sure what is happening on a micro level, but I experimented with the addition of a specific wine making yeast. It took a few failures to figure out proportion, but as soon as the flask cools I add @ 2g of the inactive yeast in sterile arena, and give them 15 min to float and do their thing. I usually run my fragmentation protocol while the yeast is activating, after adding the mother culture I start the spin, so I am running both cultures PARALLEL, then at the end I peroxidate and the yeast are destroyed. My running theory is that the myc is grabbing the yeast as well as the ME while they are also healing from the frag and training enzymes for the next substrate. I have also experimented with not killing the yeast, and injecting the parallel culture into PF style jars, PC takes forever to fruit but captures in record time. The fruits are unusually dense, have a yeast tinge where they should be white and are very heavy. I would say experientially that evidence points to an unusually active flush considering I ran same strains parallel and did a taste test side by side on week after another. I would really like to talk more about this, but the lady of the house is baking choc chip cookies just in time for munchies-- who-hoo, munchies... Mycoelf -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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Quote: I have a special incubator that uses a 15W full spectrum compact fluorescent as a heat source. The bulb is wired to a thermostat. intermittently the bulb fires and gives the light exposure required to initiate pinning on petri's, for the purpose of evaluating a strains willingness to pin. I sterilized the RCG and dropped it at the perimeter of a new but established culture. The section in the center got real thick and did not colonize further, then it exploded radially with the thinner "running" myc, and when it hit the boundary, it fruited. If you look in the pic there are many "nodules" that would give rise to many fruits under different circumstance. I.E a spawn run. what amazed me is the spores that it dropped in culture. I had never seen a sporulateing pin on a dish before that, This strain originate from a clone that had been spawned using the fermentation method. I don't know if it means anything, but i could hardly coax the myc to colonize much less run. lATER mycoelf -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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Coolaid smile Registered: 10/02/09 Posts: 897 Loc: SW WA Last seen: 11 years, 3 months |
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It is clear that you have a lab to work in unlike most of us who use our kitchen sink and counter. How big is your lab mycoelf?
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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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I have a space that is 18X 18, and 2 stories. This space is divided into a clean room that is 12X12, a spawn storage room, that is 8x8 and a fruit room that is 8X11. Th remaining room is 8x 10 and is a makeup, extraction lab, and is where I autoclave. It is my 3rd lab. My first was a spare bedroom, my second was built into a auto garage and was 11X15. I had every operation except autoclave in that room, and boy let me tell you it was tight.
My current lab is currently under construction. I finally bought property and it had the above mentioned space already built and I spent the winter framing out. I still have a bit of wiring and sheathing to do and I will be ready to work again. I have not worked in sterile culture since November and boy do I miss it, The last time spent in the old lab was a mad dash to get all of my new clones for 09' into In-Vitro, And I am happy to say that they are all looking good and happily chilling out. I am including a pic my last work space- just for prosperity. Happy belated Osatra, Mycoelf -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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Coolaid smile Registered: 10/02/09 Posts: 897 Loc: SW WA Last seen: 11 years, 3 months |
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11x15 would be a dream at this time of my life. The kitchen blows to say the least. You have flasks and a flow hood yes definitely a lab and a nice one at that. We have an offer on new house with plenty of space so hopefully our offer will be accepted and my lab will be in the works in the next few months.
Back to agar... I have been using fungi perfecti's antibiotic agar and am just about out. I have some agar that I bought at an Asian mart and have some potato flakes. I was reading about making ones own agar but I have not done it yet and was wondering if you have any pointers or should I just purchase my agar of the next meaning the kind that you just add water and autoclave. myc Ps I did the transfers you suggested and they are taking off and looking great. I took to slants and now am soaking my wood chips that I plan to inoculate her in the next day or two.
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Agent Of Chaos Registered: 06/26/09 Posts: 555 Loc: hyperspace |
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I started with FP agar- and high quality agar it is. when I first started, I was afraid to make my own because I was afraid that I would mess something up and my cultures would suffer. I was also convinced that I could not live with out the antibiotic gentymiacin sulfate. I was just expressing insecurities about my noobieness. I used AGAR AGAR from a health food store, it is with the bread making stuff for an additive to make bread moist, and went from there. I use the kelp base agar, I don't know why. One of my strains is P carulecens, incidentally I found in ancient archival material that this mushie HATES rye and will not grow on any medium containing rye. I don't remember if I called FP and asked or it was on the label, but I believe that his base is wheat and rye, explaining my poor success with that strain on FP AGAR, live and learn. FP claims that their AGAR is a superior lab grade, and I believe it to be true, but you can buy the AGAR alone outside of the premix. I found that the FP AGAR slices and dices nicely under scalpel, but resist dissolving in fermented culture. The Health food AGAR dose not pour as clear but dissolves nicely in Fermentation. The reason that this is important is if your method of delivery involves passage through a fine port, such as a needle on a syringe or a peristaltic pump. I would encourage you to experiment with your own make up and record your base ingredients and note in your system somehow which agar is which, I always pour new plates before I run out and sometimes it is helpful to have different kinds depending on the type of work that you are doing. I have a clean cooler in a sterile enviorn that I keep blank AGAR culture in so they are all in one spot when I go to work, and I avoid useing a blank the same day it is poured so I know that it is a clean plate when I start. I also use sandwich baggies to store my blanks so that nothing comes in from the outside before use. I wipe them down outside and place 2 100x15 in each bag and use as needed. I don't usually keep them in the bag after the innoc, but sometimes do as an extra measure of quarantine if I have made a clone or i am not sure of a culture's purity. I also parafilm every inoculated culture
I know that someone on this list poo-pooed peroxide in agar, but I told my mouse story previously in a recent post here it is copied If you are poring from a flask remember that the #1 vector is contact. I have a couple of silicone pot holders that I pc with the agar, one to provide grip and thermal protection from the hot flask and also to allow a firm grip lower from the lip, the other to set the flask on if for some reason I must set down he flask. This prevents rapid cooling when doing a large run. If your clean alcohol-ed gloved hand comes closer than 1 inch to the top you will probably contaminate. I set up an agar run so that once the flasks picked up it does not leave my hand until I am done. I use the stack method in GGMM, while intimidating to the novice is easy once you achieve practice. A steady hand is also important, I usually smoke a bowl while I wait for the PC to achieve Zero psi. Last year I had shut down working over the winter a couple of months and a mouse moved in in my absence. First thing after an opening deep clean I poured peroxidated agar and had fumbled the lid on one plate, so I left it out on my table overnight for "observation" The next day I came in to find the lid off the plate and the agar had bee munched by the starving mouse. Deeply curious as to the real power of peroxide I wrapped the plate and quarantined it in an incubator, expecting to find contam inside 72 hours. six weeks of observation yielded no contam whatsoever. I was astonished as I discarded the plate end quote This is my testament to the power of peroxide in AGAR. It does not survive the PC but I prepare a 10 ML graduated cylinder in foil and peroxidate 8ml in 600 ml of agar. I pour and swish to distribute throughout. I keep my peroxide in the fridge and I usually forget to let it warm up before hand so it usually goes in cold. I know that Logic says that it should breakdown but I have proved to myself that it is still active not only by the above story but also years of observation. I also noted that it seems as if you are going to inoculate your wood from AGAR?? This probably wont work if that is the case. The point of the intermediate grain master is to not only provide many points of inoculation but also to provide a nutritive base for the myc as it gains a foothold in the bulk sub. 10 to20 % by volume is a good base. Less than 10% and you run the risk of the other critters overrunning the sub or an extremely slow/partial capture. You might review GGMM for the model onto a bulk sub using grain masters, You might also check out Myc Running, there is a tec in there on how to do a dirty clone on cardboard and then a strategy for laying out outdoor beds, believe it or not this tec features the very same mushie that you are attempting to cultivate. You are looking for a 38 to 50% moisture content in your wood for colonization. I usually do this by "feel" but you could weigh a representative sample in a container at 100g, then bake at 350 for three hours and reweigh. Don't forget to tare the container. the second weight is the percentage of dry mass minus the water, then you can do the math. I could not decrypt the PM you sent me as I am not yet a proper member, could you resend your reply with no code? Good luck with your agar. Mycoelf -------------------- Mycoelf Sterility is a process that can be likened unto infinity, which is a long walk, the closer to the end you start before beginning, the more achievable the goal of infinity becomes. Remember, cleanliness in next to goddessness ![]() ![]() ![]() ![]() ![]()
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