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Offlinerandy420rhoads
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Oysters not pinning
    #14051169 - 03/01/11 09:07 PM (12 years, 10 months ago)

After a mishap with my oyster culture syring I was only able to knock up 4 jars successfully using the Pf tek on BRF( just a couple so I can clone to agar then grow in bulk). According to RR's videos you must leave them in the jars for 1 week after full colonization before birthing (or they will just sit in the FC for a week anyway) wich I did. after a little over a week in the jars after 100% I birthed, dunked for 24 hours then rolled in dry verm. I placed the cakes in idividual plastic milk jugs with 1-2 inches of perlite.


They've been in the FC for a week now with no pinning, or even mycelial knotting. They've almost covered the dry verm case in mycellium. The temps are between 60-72 with light misting and fanning 5-10 times a day. What's going on


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OfflineHumility
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Re: Oysters not pinning [Re: randy420rhoads]
    #14051340 - 03/01/11 09:38 PM (12 years, 10 months ago)

Did they smell fresh, like mushrooms when you smelled them?

You placed the cakes in individual plastic milk jugs?  Oysters are very sensitive to CO2 buildup; they need lots of fresh air all the time.  That doesn't sound like a healthy environment for them.


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: Humility]
    #14051402 - 03/01/11 09:47 PM (12 years, 10 months ago)

Yes. I grew cubensis years back, these a different yet similiar smell. A much more pleasant one distinctly healthy mushroomy without the tang of cubensis.



I know they are sensitive to CO2 buildup I open and fan at least 5 times a day mostly an upwards of 10 times a day if not more. Is that not a sufficient rate? They still smell very healthy and are colonized the rolled casing, just not pinning.


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OfflineBlueLightRain
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Re: Oysters not pinning [Re: randy420rhoads]
    #14052650 - 03/02/11 02:35 AM (12 years, 10 months ago)

Hmmmm. Pinning development is truly strain specific. For instance I have a strain of Pl. pulmonarius that pinned a couple days after I exposed it to air. I have a strain of Pl. ostreatus that doesn't fruit for a week after being exposed to fresh air. You could be close.

Also, I never misted my Pl. pulm the first time it grew (and never misted the fruit either!) and I only misted the Pl. ostreatus once to twice a day. You might be drowning it. Let it breath more and give it a bit of a break from the water. Too much water can be restricting.


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: BlueLightRain]
    #14054034 - 03/02/11 11:17 AM (12 years, 10 months ago)

It's sprinkling out i'm leaving it outside (under cover) with the lid off. Maybe It just needs air.


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: randy420rhoads]
    #14054162 - 03/02/11 11:42 AM (12 years, 10 months ago)

Well just checked the one outside. Some weird shit going on. Looks like 2 different contams dont understand how they could have done this overnight on a fully colonized cake.

One spot looks gray, the other looks like caramalized MYA. Both spots are roughly dime sized on part of it.



FInally one of my indoor cakes has 7 tine primordia forming on it wich i'm thankfull foor but why so little i thought oysters had huge numbers of pins?

Also noticed they formed on the base in between the jar lid and cake (I set the cakes on upside down jar lids w/ rings. My best guess is they fruited between there because it was more humid. Does this soudns correct?


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OfflineRogerRabbitM
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Re: Oysters not pinning [Re: randy420rhoads]
    #14054166 - 03/02/11 11:42 AM (12 years, 10 months ago)

Oysters won't pin in a milk jug.  The CO2 level is too high.  If they're non-contaminated and a viable culture, you'll see pins with them outside in the rain.
RR


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: RogerRabbit]
    #14054217 - 03/02/11 11:54 AM (12 years, 10 months ago)

Ok i'll take that info for the next batch for sure, but they did start pinning. Did you read the most recent post above yours?


And I don't understand how the Co2 level can be too high if i fan say every hour?


Edited by randy420rhoads (03/02/11 11:55 AM)


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OfflineBlueLightRain
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Re: Oysters not pinning [Re: randy420rhoads]
    #14054299 - 03/02/11 12:14 PM (12 years, 10 months ago)

Pins will develop according to how much sustenance is available to the oysters. If its half a pint of substrate, you'll probably get a small pin flush. If it's 20 pounds of substrate, well whoa momma hold on!

Oysters love fresh air...I've seen them malform in shotgun terrariums.


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: BlueLightRain]
    #14054351 - 03/02/11 12:26 PM (12 years, 10 months ago)

AS long as I can get a good tissue sample/ prints from at least one.

How do you print clone? Just like cubensis? Cut down the stipe under sterile conditions, removed some mycellium and place on agar?


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OfflineBlueLightRain
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Re: Oysters not pinning [Re: randy420rhoads]
    #14055189 - 03/02/11 03:04 PM (12 years, 10 months ago)

Yeah if you want to start your own strains from scratch, take some prints. If you want to clone exactly what has grown, remove the innards of the mushroom and transfer to agar. This comes with a hole host of difficulties. Your best bet is to make some prints. This way you can watch the mycelium on the plates and transfer the strains that grow most vigorously.


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: BlueLightRain]
    #14055510 - 03/02/11 04:10 PM (12 years, 10 months ago)

I have some agar plates i filled up last night so i'll probablly do both.

RR never got back to me about this question.... HE made it seem like you want or it's at least ok to get contams on agar, then seperate the healthy mycellium to another agar plate. Is that true?


The lids on my plate also seem pretty loose like they'd let airflow in and compromise sterility, is it normal for them to be so loose?


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OfflineBlueLightRain
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Re: Oysters not pinning [Re: randy420rhoads]
    #14056036 - 03/02/11 05:47 PM (12 years, 10 months ago)

Yeah I've found its difficult to avoid some kind of contamination when cutting out a mushroom's innards. Expect bacteria of some sort then keep transferring the fastest healthiest looking mycelium away until you get a pure culture.

I'm pretty sure all brands have loose lids, but they make a tight seal where the rim contacts the lid. Pick up some 2" Flat Twine at Ace hardware and use that to wrap your plates. I believe it's wood-based cellophane. It breathes and works the same as parafilm and its cheaper. It stretches well and clings to plastic phenomenally, as well as sticking to itself.


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: BlueLightRain]
    #14056209 - 03/02/11 06:19 PM (12 years, 10 months ago)

Appreciate all the advice!

So after I transfer my first clone expect contams and just keep transfering. Got it. Is it a bad idea to try and use some as spawn (a wedge for a rye bag in this case) before transfering and getting a completely clean plate? Or use as spawn before the plate is 100% colonized?


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OfflineBlueLightRain
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Re: Oysters not pinning [Re: randy420rhoads]
    #14058174 - 03/03/11 02:00 AM (12 years, 10 months ago)

Quote:

randy420rhoads said:
Is it a bad idea to try and use some as spawn (a wedge for a rye bag in this case) before transfering and getting a completely clean plate? Or use as spawn before the plate is 100% colonized?




Respectfully, yes. Since you're likely to have contaminants on your agar plate then you're even more likely to get contaminants on your rye grain, given that it has more surface area that it can touch. Take the time to isolate the mycelium and make it pure and you'll have an endless bounty of fungus from which to draw. If you try and jump the gun you'll have to start all over. I recommend next time you get spores from a syringe, make some drops across a bunch of plates. This way you can isolate many different strains and select those that are most vigorous.

You don't have to wait until your plate is 100% colonized to use it. In fact I recommend once it is almost completely colonized to then create more plates. Each plate can then either create more plates (we are talking exponential factors here!) or be used to create grain spawn. Think of the possibilities!


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: BlueLightRain]
    #14060284 - 03/03/11 02:00 PM (12 years, 10 months ago)

Wow so streak some spores on an agar plate with an inoculation loop rather than making syringes growing then cloning?

If I do a steak of spores wont there be dozens in not hundreds on strains on such a small surface area it would be hard to tell wich is healthier?


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OfflineBlueLightRain
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Re: Oysters not pinning [Re: randy420rhoads]
    #14064422 - 03/04/11 02:03 AM (12 years, 10 months ago)

Yeah if you make a nice solid culture bank to draw from you can do this an endless number of times. Let me do some simple math for you. Once I discovered this I was a completely changed man. You can take one of those syringes and inoculate a few jars -OR- you can drop some spores on several plates. Once you isolate a strain, you can allow it to colonize your plate, lets label this p1. Once this mycelium (p1) has colonized the plate you can then comfortably make 20 plates from little wedges you transfer to other plates (call them p2). If you then allow each of these wedges to grow out and colonize a plate, you have enough to mycelium to inoculate a bunch of jars. But you probably only need one plate to inoculate around 10 jars. You can then take the other 19 plates and make 20 more plates PER plate. So you then have 20 times 20 = 400 plates of mycelium (p3). Do this again and you now have 8000 plates (p4). In case you are worried that you will use 8000 plates in the next couple years, consider expanding each plate another 20 times. Now you have 160,000 plates (p5) of mycelium (a culture bank) from which you can inoculate your jars. Each jar can also be expanded. You don't have to wait for all 160,000 jars to grow before you start a project though! After p2 gets going you can create a cycle where you are always expanding your culture bank while simultaneously inoculating jars. With a vigorous strain, you can go from 1 plate to 160,000 plates in a month! Over 3 million plates in 5 weeks! Marvelous!

This is mind blowing and I'm always fascinated by this! This is why its better to use your spore syringes you purchased and your prints you've acquired to make agar plates first instead of using them to inoculate jars directly.

To your second question: Yes you are right. As soon as little white spots of mycelium crop up on your plate (be sure its mycelium and not a contaminant), transfer each spot to other plates. Then look for the denser, sometimes ripply shoots and cut those out and transfer. Keep doing this until each individual strain appears to have the same observable and unique features. This can take a couple of transfers to obtain a p1.

This is from dropping a spore solution onto agar. The solution ran around the edges of the plate whenever I tilted it so you see it growing pretty much everywhere on the plate. Notice the tufts that are growing out more quickly then everything else? Use those!


I simply used the sections that were showing the fastest growth.


Then I transferred all of those sections to individual plates to watch each one grow and track its rate of growth. From there I selected the ones that were most vigorous and expanded those. (I coudn't find cheap plastic petri plates at the time so I used gerber baby food jars, sterilized them then dropped the sterile agar inside...worked fine, but just gave poor visibility and little surface area. Plastic petri plates are superior)


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: BlueLightRain]
    #14072321 - 03/05/11 06:52 PM (12 years, 10 months ago)

Appreciate you taking the time to write all the good info down for me.

All of them except the one I had outside started fruiting and look quite healthy.


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OfflineBlueLightRain
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Re: Oysters not pinning [Re: randy420rhoads]
    #14075082 - 03/06/11 10:57 AM (12 years, 10 months ago)

Yay! Do you have pictures?


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Offlinerandy420rhoads
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Re: Oysters not pinning [Re: BlueLightRain]
    #14075280 - 03/06/11 11:55 AM (12 years, 10 months ago)

Seeing as you took the time to help me out so much I can definately take the time to snap a few.



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