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OfflineDeej3987
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G2G colonizing problems?
    #14231325 - 04/03/11 02:11 PM (12 years, 10 months ago)

I have successfully G2G transfered rye many times but two small problems I keep running into is a few jars not colonizing after the transfer.. The mycilium turns a yellowish color as if it dried up.

I don't know if I waited to long to break up the jars when I transfered them but I usually have about 1/4th of my jars stall out like this and I would like to eliminate this problem..

The other problem is the grains looking like they get to wet and I end up with a yellow slime where the mycilium will not colonize. I looked up contams and I'm pretty sure its mucus but I turned my incubation temperature down to 75F and I haven't had this problem as much, but it still happens.. when the temp is at 80F I notice the slime being more of a problem for me. Sometimes Ill break the jar up and the mycilium will dominate. Other times when I break it up the slime dominates... :confused:

I'm not exactly sure how to avoid this mucus typed contam? if it is mucus..

I usually break up all of my jars a couple of days before I use them again just to make sure they recolonize and I know they're good.

Any tips or advice would be greatly appreciated.


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Offlinethelivingfreekshow
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Re: G2G colonizing problems? [Re: Deej3987]
    #14231333 - 04/03/11 02:13 PM (12 years, 10 months ago)

Pictures?

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Offlinedeadmandave
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Re: G2G colonizing problems? [Re: thelivingfreekshow]
    #14231340 - 04/03/11 02:17 PM (12 years, 10 months ago)

The mucus/slime sounds just like bacterial contamination. Bacteria thrive at higher temps so it makes sense that it was winning above 75F. You can colonize at lower temps but that wont solve your problem entirely.

It sounds like you need to take a look at your procedure from start to finish and find out when and how your grains are getting contaminated. It could be the initial spawn is contammed, it could be G2G in open air or a dirty environment.

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OfflineDeej3987
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Re: G2G colonizing problems? [Re: deadmandave]
    #14231379 - 04/03/11 02:29 PM (12 years, 10 months ago)

My pressure gauge broke at the time I sterilized the rye grains.. I had to guesstimate the pressure so maybe that is the problem.. Ill just start over completely and see where that takes me.. If the problem continues Ill take pictures.. If the problem stops Ill try to get a pic on here anyways, maybe it would help others..

Do you think the dry looking Myc could be from the same bacteria?


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OfflineDeej3987
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Re: G2G colonizing problems? [Re: Deej3987]
    #14231450 - 04/03/11 02:49 PM (12 years, 10 months ago)

I found a list of all the contaminations with descriptions and pictures on here before can anyone tell me where I can find this? I'm having troubles finding it again.


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OfflineDeej3987
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Re: G2G colonizing problems? [Re: Deej3987]
    #14231917 - 04/03/11 04:27 PM (12 years, 10 months ago)

http://www.shroomery.org/forums/showflat.php/Number/14231904/vc/1#14231904

Bacteria, this is what my grains look like in some spots.. idk how to prevent this ?


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Re: G2G colonizing problems? [Re: Deej3987]
    #14232051 - 04/03/11 04:55 PM (12 years, 10 months ago)

Simply practice sterile procedures. Be sure to pc fully and work in a clean sanitary environment.

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OfflineDeej3987
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Re: G2G colonizing problems? [Re: deadmandave]
    #14234822 - 04/04/11 04:59 AM (12 years, 10 months ago)

can this happen from LC?


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Re: G2G colonizing problems? [Re: Deej3987]
    #14234877 - 04/04/11 05:44 AM (12 years, 10 months ago)

Quote:

Deej3987 said:
can this happen from LC?



It sure can. There is almost no way of knowing if an LC is contaminated, so that could very likely be the source of your bacterial contamination. And it sounds like that is exactly what you have.

As has been already said: sterilize your grains properly, inoculate in a clean environment (ie. glovebox) and use what you know to be a clean culture (NOT LC). Your problems should go away.


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Re: G2G colonizing problems? [Re: Deej3987]
    #14234879 - 04/04/11 05:45 AM (12 years, 10 months ago)

Bacteria is a huge problem when dealing with LC. This is why people go to great lengths to make fancy jar lids for their LC jars. I use a rubber gasket under the lid on mine as an extra precaution.

So I wouldn't switch to LC to avoid this problem. You're better off with g2g.

Just make sure your grain masters are clean. Start them from agar. And stop incubating unless it's cold there. Temperature has a huge affect on bacteria.


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OfflineDeej3987
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Re: G2G colonizing problems? [Re: anonjon]
    #14239965 - 04/05/11 02:07 AM (12 years, 10 months ago)

Can I get this from a spore print? How about a syringe? Ive been using the same syringe for a while.. How can I clean my syringes so I can reuse them? If I pc them they begin to melt... so maybe soak them in alcohol/bleach?

I do g2g transfers with my initial lc jar.. some jars from the g2g transfers will be successful while other jars get this bacteria problem.. so maybe my lc is ok and I'm not being as sterile later? or can the bacteria be there the whole time just waiting to spread? idk I'm at a loss here¿?

I also started from the beginning with a spore print (lc and ms are used with separate syringes) and the jars ended up getting bacterial growth also...

I guess the big question I have is can the bacteria be there the whole time, from per say a bad lc or ms print and its just a mater of time before the bacteria starts appearing?


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OfflinePrimal Call
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Re: G2G colonizing problems? [Re: Deej3987]
    #14240177 - 04/05/11 03:47 AM (12 years, 10 months ago)

Quote:

Deej3987 said:
Can I get this from a spore print? How about a syringe? Ive been using the same syringe for a while.. How can I clean my syringes so I can reuse them? If I pc them they begin to melt... so maybe soak them in alcohol/bleach?

I do g2g transfers with my initial lc jar.. some jars from the g2g transfers will be successful while other jars get this bacteria problem.. so maybe my lc is ok and I'm not being as sterile later? or can the bacteria be there the whole time just waiting to spread? idk I'm at a loss here¿?

I also started from the beginning with a spore print (lc and ms are used with separate syringes) and the jars ended up getting bacterial growth also...

I guess the big question I have is can the bacteria be there the whole time, from per say a bad lc or ms print and its just a mater of time before the bacteria starts appearing?




your last paragraph... yes. the cases where it isn't winning the race is luck of the draw, but like others pointed out, you are helping the bacteria by incubating above 75F.

you should order a syringe from a trusted vendor so you can start with what is assuredly clean or get agar and do some transfer work to clean up your current print/LC.

also, I PC my syringes at 15-17 PSI for 60-90 minutes and they haven't melted yet. sounds like you need to get a gauge replacement or different syringes.

hope this helps :peace:


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Re: G2G colonizing problems? [Re: Primal Call]
    #14244160 - 04/05/11 08:20 PM (12 years, 10 months ago)

Hmm how do you put the syringe in the PC, any specific way? bc when I did it before the needle part was crooked as if the plastic part bent a little bit.. it wasn't as straight as it was before if that makes enough sense ¿?


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OfflinePrimal Call
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Re: G2G colonizing problems? [Re: Deej3987]
    #14245703 - 04/06/11 12:58 AM (12 years, 10 months ago)

Quote:

Deej3987 said:
Hmm how do you put the syringe in the PC, any specific way? bc when I did it before the needle part was crooked as if the plastic part bent a little bit.. it wasn't as straight as it was before if that makes enough sense ¿?




For GLC, I aspirate full to empty 2-3 times to rinse it out then fill it again and leave it full. put the cap on the needle loosely and place in a qt jar tip up with a hemispheric foil lid (the full syringe is taller than the qt jar) to be PC'd. If you leave the tip facing down, you'll lose some of the water during PC.

For LC, same thing as above but just don't fill it with water pre-PC.


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Re: G2G colonizing problems? [Re: Primal Call]
    #14264030 - 04/09/11 06:47 PM (12 years, 10 months ago)

I will have to sterilize my syringes then thanks for the advice.

as far as sterilizing the rye grains, I usually do a 50 lbs bag at a time that way I have around 144 quart jars ready to be used.. I do the 24 hour soak, then get a barrel of water to boil, put all the rye in the barrel for 10 minutes then I throw all of the rye in a giant colander.. Now this is something I'm wondering, I let it dry for about 2-4 hours before its actually ready because there is so much.. then I load up the jars 2/3 way full and pressure cook 90 minutes at 15PSI..

  Does anyone think that me letting the rye grains sit out with a fan blowing on it for about 4 hours will allow any kind of contam such as bacteria to get on the grains and mess with it later even after I pressure cook it? I mean there is no signs of bacteria until after I've used the grain jars by g2g transfers or inoculations..


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OfflinePrimal Call
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Re: G2G colonizing problems? [Re: Deej3987]
    #14267211 - 04/10/11 01:52 PM (12 years, 10 months ago)

of course contams get on them during drying. but it doesn't matter because of the subsequent PC'ing. If you are getting contams after inoc or G2G, it could be a lot of variables, but they are obviously related to your sterile technique or your lid setup.

do you use a still air box or GB?
clean all surfaces and air?
wear gloves wiped with rubbing alcohol?
flame the needle tip for inoculations?
surgical mask?
lid setup?


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Re: G2G colonizing problems? [Re: Primal Call]
    #14267865 - 04/10/11 04:39 PM (12 years, 10 months ago)

How long does it take to get through fifty lbs of rye? If it's longer than two weeks this could be a vector of contamination

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Re: G2G colonizing problems? [Re: deadmandave]
    #14267972 - 04/10/11 05:05 PM (12 years, 10 months ago)

I find that using a cheaply made still air box made out of clear tubberware helps ALOT with contam rates. Just cut 2 holes in the box with for your arms, and wipe in out with alcohol before each use

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Re: G2G colonizing problems? [Re: Primal Call]
    #14278314 - 04/12/11 02:25 PM (12 years, 10 months ago)

Quote:

Ryath said:
of course contams get on them during drying. but it doesn't matter because of the subsequent PC'ing. If you are getting contams after inoc or G2G, it could be a lot of variables, but they are obviously related to your sterile technique or your lid setup.

do you use a still air box or GB?
clean all surfaces and air?
wear gloves wiped with rubbing alcohol?
flame the needle tip for inoculations?
surgical mask?
lid setup?




what is a still air box? I use a glovebox
I clean all surfaces and air yes.
yup I have aqua gloves.
surgical mask yes.
lid? uhh the lid is on top I'm going to make it on the side just encase contaminates are falling in it from the top.. the GB is made of plexiglass.. actually its really big  4'Long X 2' Deep X 2' Tall... I am in the process of cutting it down in size right now.
and I do not flame sterilize the needle.. I know you are suppose to but my first needle I did this with got rusty and I had to throw it away so ive just been wiping it with alcohol thoroughly.. Maybe this is my problem? I'm going to pc after a few uses for now on and maybe that will stop the bacterial growth..


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Re: G2G colonizing problems? [Re: Deej3987]
    #14278353 - 04/12/11 02:36 PM (12 years, 10 months ago)

Quote:

I do not flame sterilize the needle..




This is absolutely part of the problem. :super:

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