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Offlinewideweb
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Is this normal?
    #3364107 - 11/15/04 03:21 PM (12 years, 23 days ago)

I am making tissue culture of gills of some oyster from the grocery store for the first time. After two weeks, at 72F, there is white stuff around the tissue:



When I place the dish between the camera and a light source, I can notice mycelium:



Since I have never done this before, I am anxious to know: is this white stuff normal?

Thanks.


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Offlineragadinks
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Re: Is this normal? [Re: wideweb]
    #3364227 - 11/15/04 03:47 PM (12 years, 23 days ago)

For me that looks like a kind of bacterial contamination.
I am sure the guys in the contamination forum now better ...


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InvisibleATWAR
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Re: Is this normal? [Re: ragadinks]
    #3364782 - 11/15/04 05:42 PM (12 years, 23 days ago)

That definetly does not look like oyster mycelium to me (except the 2nd picture could show some mycelium crawling out). It looks like bacteria. If it is oyster mycelium it should cover the plate in a week, so keep an eye on it. Try a slice from inside the mushroom rather than the gills next time. It should take within a few days rather than a few weeks...


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InvisibleKoala Koolio
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Re: Is this normal? [Re: ATWAR]
    #3364991 - 11/15/04 06:35 PM (12 years, 23 days ago)

I too recently tried this with a crimini and oyster. the crimini seemed to degrade almost insantly, into brown. It was perfectly fresh, the mushroom was split open, not cut, and a piece taken from the middle. Nice and fluffy and white, for a few hours.

Are the chances of success with this method really so unlikely?


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InvisibleATWAR
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Re: Is this normal? [Re: Koala Koolio]
    #3365050 - 11/15/04 06:50 PM (12 years, 23 days ago)

Quote:

elgr said:
Are the chances of success with this method really so unlikely?





Not at all. My Pleurotus citrinopileatus culture came from a grocery store clone. It has been done by several members with more than just oysters. It does indeed work, although you must remember that the cultures you obtain through this method have gone through an unkown number of transfers (cell divisions). What you end up with may not be able to be expanded very far, or fruit very well at all (my golden oyster is no longer golden, but rather "tinted" or "faded" instead). Not to mention that there is no way to know to what substrate or what conditions the strain is adapted to...


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InvisibleKoala Koolio
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Re: Is this normal? [Re: ATWAR]
    #3365323 - 11/15/04 07:31 PM (12 years, 23 days ago)

Good points. But any idea why mine seems to be failing? air exchange perhaps...


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InvisibleATWAR
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Re: Is this normal? [Re: Koala Koolio]
    #3365801 - 11/15/04 08:48 PM (12 years, 23 days ago)

Could be air exchange problems, I dunno. Sounds like contamination to me. You say it degraded into "brown"... Huh? You mean it turned to liquid? If so, that would indicate either poor air exchange, bacteria, or both. It could also be due to no so fresh mushrooms being used...


I would try again as soon as a fresh mushroom supply hits the store. I am sure you could ask them when that happens. Also, grocery stores have a policy of "rotating stock" where the new shipments will be placed behind the old ones on the shelf. Look through all the mushroom containers and pick the fattest, freshest, healthiest looking oyster you can...


I have never had a problem with air exchange in my petris, I seal them up with electrical tape (I suspect it allows at least some air exchange, as I can smell the mycelium through it). You could try getting some of the porous medical tape if you so desire. You could also try a dip into peroxide solution (0.3%) before placing the excised tissue onto the plate. I do this most of the time as it helps to hydrate the tissue, and kill most of the nasties.


Good luck!


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Offlinefalcon
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Re: Is this normal? [Re: Koala Koolio]
    #3371563 - 11/16/04 11:27 PM (12 years, 22 days ago)

Try starting more than one plate.
Give it some time, ussually discoloring is not a good
sign, sometimes they recover. Crimini's will suck up moisture
from the agar, become waterlogged and so change
color.


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InvisibleKoala Koolio
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Re: Is this normal? [Re: falcon]
    #3371776 - 11/17/04 12:20 AM (12 years, 22 days ago)

"Crimini's will suck up moisture
from the agar, become waterlogged and so change
color. "

This seems to be exactly what happened. I know what most contams look like, and it doesn't appear to be one. Its definately not spreading, and doesn't appear to be on top of the agar, its just a color change possibly because like oyu said its sucking up moisture, it looks incredibly wet, and theres absolutely no air exchange.

On the bright side, the crimini is now covered in a nice fluffy white mycellium. I guess it just takes some patience :smile: just to be safe, I'm going to transfer it to a few other plates, see what happens.

Oyster still hasn't done much. Damn sample went directly to the side of the dish. Tried shaking it around but it simply clung to the side and moved around. oh well. theres always the chance to do it again.

Have never worked via cloning a fruit, but I'm successfully germinated spores on agar, and isolated substrains and grown them out.


--------------------
You're not like the others. You like the same things I do. Wax paper, boiled football leather... dog breath. We're not hitch-hiking anymore, we're riding!


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Offlinefalcon
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Re: Is this normal? [Re: Koala Koolio]
    #3376477 - 11/17/04 10:45 PM (12 years, 21 days ago)

On the bright side, the crimini is now covered in
a nice fluffy white mycellium.
  :thumbup:
sounds like it's happy keep us posted on how its doing.

Give the oyster another try sometimes it takes a
couple of tries to get a good culture going.


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InvisibleKoala Koolio
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Re: Is this normal? [Re: falcon]
    #3377016 - 11/18/04 12:45 AM (12 years, 21 days ago)

Its been a day since it started, I know its not long, but its covering the mushroom bit. However, it hasn't taken off on the agar yet. I had a number of mexicana transfers onto this same batch. One of them decided it would do the same thing. Turned very fluffy (on a slice that was identical, or should be) but didn't spread into it. I knocked the wedge against the side, and it even started growing on the glass?

Something seems quite wrong.


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You're not like the others. You like the same things I do. Wax paper, boiled football leather... dog breath. We're not hitch-hiking anymore, we're riding!


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Offlinefalcon
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Re: Is this normal? [Re: Koala Koolio]
    #3381955 - 11/18/04 11:49 PM (12 years, 20 days ago)

I knocked the wedge against the side, and it even started growing on the glass?

That doesn't seem unusual.


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InvisibleKoala Koolio
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Re: Is this normal? [Re: falcon]
    #3382150 - 11/19/04 12:36 AM (12 years, 20 days ago)

My new theory is moisture. I don't know why I didn't think of it earlier. These dishes were made a few weeks ago. The mexicana isolates I used on them were all within a few days of each other, however, it *was* the last one that didn't work as well. Now this crimini is doing quite the same.

And it probably liked the moisture on the side, as well as everything the shroom bit had absorbed. Any way to moisten it with sterile water that would be recommended? If so, just a couple drops I'd assume?

Sorry for hijacking the thread.


--------------------
You're not like the others. You like the same things I do. Wax paper, boiled football leather... dog breath. We're not hitch-hiking anymore, we're riding!


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Offlinefalcon
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Re: Is this normal? [Re: Koala Koolio]
    #3388281 - 11/20/04 10:08 AM (12 years, 19 days ago)

There should be enough moisture in the agar. I don't
think you would have to add any water.


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OfflineLactarius
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Re: Is this normal? [Re: Koala Koolio]
    #5199181 - 01/19/06 04:54 PM (10 years, 10 months ago)

Also tried my hand at tissue culture for the first time. Tried A. Bisporus to test my success rate while I look for some Oysters.  Transferred nice thin stringy tissue from middle of stem to PDA in honey jar.  Sealed up with masking tape.  It is 24hrs later and most are brown, dont know if others will succeed.  :frown:  Tried some tissue to Dextrose Jar also tonight.

Have tried transferred some fresh Camembert chees rind to PDA.  Looks good, althought it is growing slow.  Think my house is too warm now.  27-30 degC during the day.  How does penicillin Camemberti fruit....


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Mushrooms, Mycology and Psychedelics >> Gourmet and Medicinal Mushrooms

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