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OfflineBlutripper
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Inoculation/colonization problem
    #19464685 - 01/23/14 08:36 PM (10 years, 11 months ago)

I'm a first timer, working with pf steam tek...brown rice flour/vermiculite mix....I steam my jars and wait 24 hours to inoculate....and here's my problem...after I inoculate, in most of the jars nothing grows... A few jars will start colonizing at only a single inoculation point...it takes so long for them to colonize after a month and a half I usually get some contamination....is there something I am doing wrong or more likely are the spore syringes bad?

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Re: Inoculation/colonization problem [Re: Blutripper]
    #19464823 - 01/23/14 09:02 PM (10 years, 11 months ago)

Walk me step by step exactly what you do from mixing the BRF to setting the jars on a shelf.

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OfflineRogerRabbitM
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19464854 - 01/23/14 09:08 PM (10 years, 11 months ago)

More than likely too wet, but you'll have to explain your processes or nobody can help.
RR


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OfflineBlutripper
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19479427 - 01/26/14 11:01 PM (10 years, 11 months ago)

Ok, this is my process. 

Jar lids prepared with 4 holes in each.

In a bowl I put 2 cups of vermiculite and then add 1 cup of water, mix them up well.  Then add 1 cup BRF and mix that in.  Use a spoon to put the mixture into 5 jars leaving a half inch or so empty at the top of each jar.  I use first a dry paper towel to clean the rim of the jar then a wet paper towel.  Then I use a spoon to scoop dry vermiculite into the top of each jar.  Put the lids on the jars, rubber side down and Cover the tops in tin foil.

I use a pot with a good lid to sterilize.  The first time I used three lid rings on the bottom of the pot and a layer of tin foil to keep the jars off the bottom.  The second time I added a second layer of lid rings and tin foil.  The first time I filled the pot with water until it came just over the tin foil and the jars were sitting in less than 1/4 inch of water...the second time I kept the water just below the tin foil layer so the jars were not in contact with the water.(i changed tactics as i thought perhaps i had steamed water out of the jars by getting them to hot). I could sterilize three jars at a time and did 12 the first time and 10 the second. So it took a while each time.    I set the burner at the lowest setting which kept the water simmering.  After the water started bubbling for each set of three I would wait 60-75 minutes then remove the pot and set it on a towel for 10-15 minutes before removing the jars and also setting them on a towel to cool then start the next set if three...

I would then wait 24 hours and remove the tin foil.  Most of the jars had a little bit of moisture under the tin foil that had seemed to have had steamed out.  But it was very little.  I would then use a purchased spore syringe to inoculate each of the four holes in each jar with 1-2 cc's.  I was sure to tap the syringe on the sides and shake it before beginning. 

I will add I ordered the syringes the first week of November.  While there was no visible damage it's possible the experienced near freezing temps in transit. 

After I finished inoculating the jars I put them in the cupboard and would check them every four days or so. 

Most of the jars had nothing happen.  A few had one inoculation site start colonizing and one had two.  Of the ones which did start to colonize Most took a couple of weeks to even start. Of those only one fully colonized and is currently fruiting nicely.  Most would show contamination after a month to a month and half, and a few just stalled out and the mycillium quit growing.

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OfflineKizzle
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19480110 - 01/27/14 05:21 AM (10 years, 11 months ago)

What kind of contamination?


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OfflineBlutripper
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Re: Inoculation/colonization problem [Re: Kizzle]
    #19483731 - 01/27/14 09:33 PM (10 years, 11 months ago)

A few get a black mold starting at the top... Others get a grey mold at the inoculation site.... But that's not really what I'm worried about as it happens after a month and half -two months...I'm more wondering why the spores won't colonize...

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Invisiblejenkinz
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19484028 - 01/27/14 10:48 PM (10 years, 11 months ago)

Did you flame sterilize the needle between jars? Are you inoculating in a still air box, or open air? Did you inoculate up against the glass? Did you buy from a reputable vender?


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OfflineKizzle
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19484095 - 01/27/14 11:07 PM (10 years, 11 months ago)

Usually when there's no growth after the inoculation it's contaminated with bacteria.

You mentioned mold growing near the dry verm layer in some, it's possible you're dry verm layer is getting wet which allows bacteria or in some cases mold to colonize through it.


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Invisiblejenkinz
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Re: Inoculation/colonization problem [Re: Kizzle]
    #19484136 - 01/27/14 11:19 PM (10 years, 11 months ago)

OH! Do you put tape over the holes in the lid, THEN put foil on it? If not, the steam may be getting in and wetting the dry verm layer.


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Offlinevikingsc
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Re: Inoculation/colonization problem [Re: jenkinz]
    #19484208 - 01/27/14 11:37 PM (10 years, 11 months ago)

Quote:

jenkinz said:
OH! Do you put tape over the holes in the lid, THEN put foil on it? If not, the steam may be getting in and wetting the dry verm layer.



I've had that fuck me up several times.

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Invisiblejenkinz
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Re: Inoculation/colonization problem [Re: vikingsc]
    #19484234 - 01/27/14 11:44 PM (10 years, 11 months ago)

Me, too, buddy. Me, too.


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OfflineBlutripper
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Re: Inoculation/colonization problem [Re: jenkinz]
    #19488668 - 01/28/14 09:09 PM (10 years, 11 months ago)

I did not flame sterilize the the needle...open air....up against the glass...not sure about the vendor(that's part of what I'm trying to figure out in posting)...

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OfflineBlutripper
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Re: Inoculation/colonization problem [Re: jenkinz]
    #19488671 - 01/28/14 09:10 PM (10 years, 11 months ago)

I did not put tape down...what sort of tape would one use?

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OfflineBlutripper
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Re: Inoculation/colonization problem [Re: Kizzle]
    #19488678 - 01/28/14 09:11 PM (10 years, 11 months ago)

So if it contaminated with bacteria how do I solve this?...longer sterilization?

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Offlinemycopanda
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19488717 - 01/28/14 09:20 PM (10 years, 11 months ago)

Quote:

Blutripper said:
I did not flame sterilize the the needle...open air....up against the glass...not sure about the vendor(that's part of what I'm trying to figure out in posting)...



This is a cluster fuck of a example of what your issue most likely is. unsterile inoculation, you probably had a contaminant from the moment you put the needle in. You just don't visibly see any until a month or two..


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Offlinevikingsc
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19488721 - 01/28/14 09:20 PM (10 years, 11 months ago)

Quote:

Blutripper said:
I did not put tape down...what sort of tape would one use?



As long as you take the tape off at noc time you can just use scotch tape. The only tape you can have on after noccing up is 3m micropore tape. You should always flame sterilize your needle before every jar. If you don't have an alcolohol lamp a big lighter will do. An alcohol lamp is so much faster to get red hot and you can do it with one hand. An alcohol lamp is probably the best $5 investment you can make in this hobby after maybe a SAB. I've found them in thrift stores for like $1 or $2.

If your having major contam issues it never hurts to sterilize for longer. Are PC'ing or steam sterilizing your jars?

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Offlinemycopanda
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Re: Inoculation/colonization problem [Re: vikingsc]
    #19488744 - 01/28/14 09:25 PM (10 years, 11 months ago)

Quote:

vikingsc said:
Quote:

Blutripper said:
I did not put tape down...what sort of tape would one use?



As long as you take the tape off at noc time you can just use scotch tape. The only tape you can have on after noccing up is 3m micropore tape. You should always flame sterilize your needle before every jar. If you don't have an alcolohol lamp a big lighter will do. An alcohol lamp is so much faster to get red hot and you can do it with one hand. An alcohol lamp is probably the best $5 investment you can make in this hobby after maybe a SAB. I've found them in thrift stores for like $1 or $2.

If your having major contam issues it never hurts to sterilize for longer. Are PC'ing or steam sterilizing your jars?



I believe he said steam sterilizing..

Oh and hey!! :hi:


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Offlinevikingsc
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Re: Inoculation/colonization problem [Re: mycopanda]
    #19488856 - 01/28/14 09:46 PM (10 years, 11 months ago)


Quote:

mycopanda said:
Quote:

vikingsc said:
Quote:

Blutripper said:
I did not put tape down...what sort of tape would one use?



As long as you take the tape off at noc time you can just use scotch tape. The only tape you can have on after noccing up is 3m micropore tape. You should always flame sterilize your needle before every jar. If you don't have an alcolohol lamp a big lighter will do. An alcohol lamp is so much faster to get red hot and you can do it with one hand. An alcohol lamp is probably the best $5 investment you can make in this hobby after maybe a SAB. I've found them in thrift stores for like $1 or $2.

If your having major contam issues it never hurts to sterilize for longer. Are PC'ing or steam sterilizing your jars?



I believe he said steam sterilizing..

Oh and hey!! :hi:



Hahaha :hi:
Ahh ya so he did 60-75 min steam sterilizing. 60 minutes isn't long enough for steam sterilizing. That's long enough for PC'ing. Steam sterilizing a PF jar you need to steam sterilize somewhere between 90 - 120 minutes after after a full boil. I recommend 120. Many get away with 90.

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Offlinemycopanda
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Re: Inoculation/colonization problem [Re: vikingsc]
    #19488895 - 01/28/14 09:56 PM (10 years, 11 months ago)

:whathesaid::hug:
Quote:

vikingsc said:

Quote:

mycopanda said:
Quote:

vikingsc said:
Quote:

Blutripper said:
I did not put tape down...what sort of tape would one use?



As long as you take the tape off at noc time you can just use scotch tape. The only tape you can have on after noccing up is 3m micropore tape. You should always flame sterilize your needle before every jar. If you don't have an alcolohol lamp a big lighter will do. An alcohol lamp is so much faster to get red hot and you can do it with one hand. An alcohol lamp is probably the best $5 investment you can make in this hobby after maybe a SAB. I've found them in thrift stores for like $1 or $2.

If your having major contam issues it never hurts to sterilize for longer. Are PC'ing or steam sterilizing your jars?



I believe he said steam sterilizing..

Oh and hey!! :hi:



Hahaha :hi:
Ahh ya so he did 60-75 min steam sterilizing. 60 minutes isn't long enough for steam sterilizing. That's long enough for PC'ing. Steam sterilizing a PF jar you need to steam sterilize somewhere between 90 - 120 minutes after after a full boil. I recommend 120. Many get away with 90.




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OfflineBlutripper
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Re: Inoculation/colonization problem [Re: Blutripper]
    #19493344 - 01/29/14 07:35 PM (10 years, 11 months ago)

Thanks for all the advice and input every one....I really appreciate it!!!....

Though I do have one follow up....I'd say I had almost as many jars do nothing at all...no colonization or noticeable contamination.... Just nothing....prolly as many jars did this as got contamination if not more.... What are your thought on why that happened?....bad spore syringe?

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