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Unble to get P. Cubensis Spores to Germinate on Agar.
    #16380326 -

Gotten MANY different varieties and species of mushroom spores to germinate onto several different agar recipes, varying the amounts of ingredients with success except for cubensis.  I can't figure out why I can't get cubensis spores to germ. on agar.  Never played with changing the water amounts per recipe though.


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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: hamloaf] * 2
    #16380830 -

Use half to 2/3 the amount of agar-agar in the recipe so it's more moist.  Dip your inoculating loop in the agar to cool it after flame sterilizing, and then take the agar covered loop to grab your spores.  This ensures they're partly covered in the agar to keep them moist while they germinate.
RR


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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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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: RogerRabbit]
    #16380910 -

How old werethe spores?  I had a print that was a year old and it took over 3 weeks to notice any growth.  Didnt help they got suspended in the gel but everything went fine afterwards.


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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: runningfox2002]
    #16381798 -

The spores are/were fresh, taken just a few months ago, independently.  The same spores exploded on sterilized grain mediums (WBS) with in five-six days. 

RR, the freshly heated, flame sterilized inoculation loop is cooled in the sterilized agar medium before contact with the spores are made, as you described.  Haven't ever tried using half to 2/3 the amount of agar-agar the recipe usually calls for though.  I'll try that the next time I make agar and let you know how it goes.


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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: hamloaf]
    #16381813 -

This is weird Ham....something unlikely happened...you know your stuff
and so this seems odd to me.

Use RRs trick to help hydrate the spores.  This ought to do.

I am not thinking that anything as complex as a water-filled syringe,
under pressure, should be necessary for fresh cube spores.

Hrmmmm

JD


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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: Javadog]
    #16381895 -

Yeah, when ever I cool the freshly flame sterilized inoculation loop in the freshly sterilized agar medium, agar medium residue coming up with the inoculation loop is an affect of doing that.  Happens every time.  I'm definitely inclined to believe that it's a spore moisture issue so, I think that using half to two-thirds the amount of agar-agar the recipe I'm using called for will do the trick.  It's the only thing described above that I haven't thought of or tried.  I'm going to try that the next time I make agar and I'll let you know how it goes.


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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: hamloaf]
    #16382100 -

Right on.

Good luck.

A fallback might be to first make a spore syringe and let it sit
for a few days before using it, and then the syringe with a weight or
rubber band to maintain higher pressure during the soak.

Let us know,

JD


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Boyd Rice told my brother that life is a corny pack of freesakes

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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: Javadog]
    #16384064 -

Right on yeah, I'm familiar with the forced hydration of spores with a syringe and a rubber-band trick.  I don't think forced hydration of the spores is necessary in my case because the spores are fresh.  I think the spores are becoming dehydrated after they've been placed on the freshly sterilized agar medium due to my agar being to hard and dry.  It makes sense to me that a moister, softer agar mix would allow for more moisture evaporation at the medium's surface, creating a more humid micro climate for longer that would keep the spores hydrated until they find mates.  This is a problem I face only with cubesis spores.  Whether taken my self or, purchased from a vendor.


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How I Get Stuff done. - My Reference Guide. - My Grows.

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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: hamloaf]
    #16384142 -

Quote:
due to my agar being to hard and dry.


That's it for sure.  An alternative is to have some sterilized water in a syringe and after swiping the spores on the plate, place a drop or two of the sterile water over the spores.  You could even do that on plates you have working now that wouldn't germinate.

I don't like spore syringes for agar work because they tend to favor bacteria for some reason.
RR


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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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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: RogerRabbit]
    #16389111 -

Haha, cool.  Yeah, all the plates I had that wouldn't germinate sat for so long, they began to grow the bacterial and mold spores that had not been killed yet, were knocked way back from sterilization.

I would speculate that just merely based on the the extra steps involved in making syringes alone is where the extra vector(s) for contamination become introduced.  You know how it is, the more you handle something, the wider the window for error becomes opened.  Why make spore syringes and then, go to agar anyway when you can just go directly from spore print to agar?  Saves time, money and, materials going from spore print to agar.


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How I Get Stuff done. - My Reference Guide. - My Grows.

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Re: Unble to get P. Cubensis Spores to Germinate on Agar. [Re: hamloaf]
    #16389782 -

Quote:
Why make spore syringes and then, go to agar anyway when you can just go directly from spore print to agar?


I would not, except to test the syringe before I used it on cakes or some such.


Quote:
Saves time, money and, materials going from spore print to agar.


Wisdom.


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Boyd Rice told my brother that life is a corny pack of freesakes

Myco-tek.org

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