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OfflineBiskop
Fokken Prawns
Male


Registered: 12/09/17
Posts: 192
Loc: Vietnam 🇻🇳
Last seen: 6 days, 14 hours
LC - No Growth After 9 Days (Josex’s Poke)
    #27472495 - 09/18/21 03:40 AM (2 months, 19 days ago)

Greetings,

Fellow Devotees of the Mush Cult,

I inoculated 3 jars of rice water with a bit of honey and agar 9 days ago. Well, I think I inoculated them. Since then there hasn’t been any signs of growth. I’m thinking the needle was probably still too hot when I did the poke and cooked the microscopic bits of mycelium. I did cool it by spraying some water into a tissue but I guess it wasn’t enough.

Is 9 days long enough to chuck it out and start over or is this normal and I need to be more patient? I’ve been watching them everyday and I’m staring to loose hope. :sad:

The LC


Thanks guys.


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OfflineJXAllen
Cannalyst


Registered: 08/06/21
Posts: 171
Last seen: 9 hours, 50 minutes
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop] * 1
    #27472511 - 09/18/21 04:11 AM (2 months, 19 days ago)

I haven't done a poke to LC broth yet. I have done LC syringe to LC broth. I added 1ml of LC from a syringe I received into a 16oz mason jar with 300ml LC broth. I used 1g LME, 1g Dextrose and 1g honey to 500ml water. I could see very small pieces of mycelium on initial inoculation. Within 7 days, the jar was half full of mycelium. I moved them to the fridge. If you're not seeing anything in 9 days, I'd say you probably didn't get a good sample.


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OfflinePBJ710
Strangler

Registered: 07/05/19
Posts: 1,062
Last seen: 2 hours, 18 minutes
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop]
    #27472513 - 09/18/21 04:13 AM (2 months, 19 days ago)

You should see growth by now if it's alive.  I'd toss it and start a new one.


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OfflineInsultingLizard
Stranger

Registered: 01/04/20
Posts: 530
Last seen: 6 hours, 41 minutes
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop]
    #27472522 - 09/18/21 04:29 AM (2 months, 19 days ago)

Just how much honey did you use? That looks super loaded. You should be using 3-4% honey per volume. It's better to err on the low side. Too much sugar is toxic to microorganisms.


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OfflineGuerrilla
Monanacahuia


Registered: 01/30/21
Posts: 1,082
Loc: United Kingdom Flag
Last seen: 14 minutes, 38 seconds
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: InsultingLizard]
    #27472525 - 09/18/21 04:31 AM (2 months, 19 days ago)

You sterilised your syringe, then wiped it with something dirty?


--------------------


Yup it’s me again. Shit, I post too much..

fahtster said:
The only bad experience is the one not had.


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OfflineBiskop
Fokken Prawns
Male


Registered: 12/09/17
Posts: 192
Loc: Vietnam 🇻🇳
Last seen: 6 days, 14 hours
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Guerrilla]
    #27472536 - 09/18/21 05:05 AM (2 months, 19 days ago)

Just a teaspoon of honey for like 500ml I think. This was my first time making an LC broth, I’ll definitely take better measurements next time. I could go only honey water for the next batch right? I like the clear jars more so you could actually see what’s going on inside. Like JXAllen’s jar. (The one I uploaded was the clearest looking of them all)

Thanks for the quick responses. I’ll throw them out and start over. Now that I know a couple more things I’ll be making some sweet LC soon. 

:cheers:

@Guerrilla, I know I’m a noob but I’m not that much of a noob haha :grin:


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OfflineGuerrilla
Monanacahuia


Registered: 01/30/21
Posts: 1,082
Loc: United Kingdom Flag
Last seen: 14 minutes, 38 seconds
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop] * 1
    #27472538 - 09/18/21 05:16 AM (2 months, 19 days ago)

Never mind I mis-read it, my bad.


--------------------


Yup it’s me again. Shit, I post too much..

fahtster said:
The only bad experience is the one not had.


Edited by Guerrilla (09/18/21 05:17 AM)


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OfflineJXAllen
Cannalyst


Registered: 08/06/21
Posts: 171
Last seen: 9 hours, 50 minutes
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop]
    #27472545 - 09/18/21 05:48 AM (2 months, 19 days ago)

I'm also using RO water.  I've seen several posts in other threads talking about using distilled/RO vs tap water. The minerals in tap water can react with the sugars and turn the broth dark.  It shouldn't be a problem, but if you want clear, use distilled or RO water.


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OfflineBiskop
Fokken Prawns
Male


Registered: 12/09/17
Posts: 192
Loc: Vietnam 🇻🇳
Last seen: 6 days, 14 hours
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: InsultingLizard]
    #27472603 - 09/18/21 08:01 AM (2 months, 19 days ago)

Ok cool I’ll just filtered water at home. A little darker shouldn’t be a problem. The jars a have now are…

Quote:

InsultingLizard said:

super loaded.



and thick. Thanks for the tip though.

Are the jars supposed to be half full or does it not matter. Most pics of LC I’ve seen are like yours or even less full.


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OfflineJXAllen
Cannalyst


Registered: 08/06/21
Posts: 171
Last seen: 9 hours, 50 minutes
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop] * 1
    #27472609 - 09/18/21 08:16 AM (2 months, 19 days ago)

You want enough head space to allow for agitating the LC without splashing whatever you're using as a filter on your jar lids. I'm using magnetic stir bars and a stir plate, so I could have filled them more. But, if you're using marbles/coins/glass/etc..  and you need to swirl to agitate, you may want a good amount of head space.


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OfflineGuerrilla
Monanacahuia


Registered: 01/30/21
Posts: 1,082
Loc: United Kingdom Flag
Last seen: 14 minutes, 38 seconds
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: JXAllen]
    #27478123 - 09/22/21 01:09 PM (2 months, 15 days ago)

I have had the exact same issue as you with Josex Poke. Only one of 5 has colonised and is looking ready for use, the only clone of the 5. I’m betting the rest won’t do anything as they’ve had long enough now.

Plenty of headroom. Far less nutrition (optimal amount of LME)


--------------------


Yup it’s me again. Shit, I post too much..

fahtster said:
The only bad experience is the one not had.


Edited by Guerrilla (09/22/21 01:09 PM)


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OfflineIngushkaya
Stranger


Registered: 07/03/21
Posts: 17
Last seen: 2 months, 14 days
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop]
    #27478132 - 09/22/21 01:14 PM (2 months, 15 days ago)

I would chuck it, it should've showed some growth by now. Sorry about your loss. :/


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OfflineBiskop
Fokken Prawns
Male


Registered: 12/09/17
Posts: 192
Loc: Vietnam 🇻🇳
Last seen: 6 days, 14 hours
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Ingushkaya]
    #27478905 - 09/23/21 12:49 AM (2 months, 14 days ago)

@Guerrilla

What do you think went wrong? I’m pretty sure my needle was too hot and cooked the tiny bit of myc, but not 100% sure because I have no experience. I’m planning on doing my second attempt today so would like to hear your thoughts.


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OfflineGuerrilla
Monanacahuia


Registered: 01/30/21
Posts: 1,082
Loc: United Kingdom Flag
Last seen: 14 minutes, 38 seconds
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop]
    #27478951 - 09/23/21 01:52 AM (2 months, 14 days ago)

I couldn’t say for sure, and you could well be right, although I do remember taking 2 samples from one plate, one right after the other, jab then jab. I can’t remember what strain but it may have been the one that grew. Give it a try


--------------------


Yup it’s me again. Shit, I post too much..

fahtster said:
The only bad experience is the one not had.


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OfflineJXAllen
Cannalyst


Registered: 08/06/21
Posts: 171
Last seen: 9 hours, 50 minutes
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Guerrilla]
    #27478990 - 09/23/21 03:16 AM (2 months, 14 days ago)

What gauge needles are you guys using? I was planning on making a LC from a couple of agar cultures. I've got 18g. I was trying to find 16g or even 14g, as the larger diameter would allow for a larger biopsy, but all I could find were blunt, liquid transfer needles. I suppose you don't need an actual needle to take a biopsy from agar though. And if you lift the lid rather than going through a self healing injection port, it wouldn't make any difference.


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OfflineBiskop
Fokken Prawns
Male


Registered: 12/09/17
Posts: 192
Loc: Vietnam 🇻🇳
Last seen: 6 days, 14 hours
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Guerrilla]
    #27478998 - 09/23/21 04:06 AM (2 months, 14 days ago)

Quote:

Guerrilla said:
I couldn’t say for sure, and you could well be right, although I do remember taking 2 samples from one plate, one right after the other, jab then jab. I can’t remember what strain but it may have been the one that grew. Give it a try




Yeah the TEK said don’t need to poke twice but it may be useful if you’re having problems. I’ll make sure the needle has cooled and then poke the agar 2-3 times just to be safe. I’ve also go 5 jars so let’s see how it goes.

@JXAllen I just checked my needles they’re 23 and 25g. I never really cared about it’s just what they sold at the pharmacy. Couple of cents each.


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OfflineGuerrilla
Monanacahuia


Registered: 01/30/21
Posts: 1,082
Loc: United Kingdom Flag
Last seen: 14 minutes, 38 seconds
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop]
    #27479018 - 09/23/21 05:26 AM (2 months, 14 days ago)

Yeah I can see it working for sure. The first jab would cool that section of the needle further, both outside and inside.


--------------------


Yup it’s me again. Shit, I post too much..

fahtster said:
The only bad experience is the one not had.


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OfflineJXAllen
Cannalyst


Registered: 08/06/21
Posts: 171
Last seen: 9 hours, 50 minutes
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Guerrilla]
    #27501533 - 10/12/21 04:14 AM (1 month, 26 days ago)

Hey guys, so I decided to take a shot at this with the agar I'm currently working with. I made two 300ml LCs in pint mason jars. I used a 18g needle for the poke, I used a fresh needle for each poke, so no flame was used. I poked both plates twice, because the sample stayed on the plate on the first poke both times.



In the AA+ jar, I could see the sample I took. It was a tiny speck of mycelium with a bit of agar attached. I could see new growth within 3 days. I gave it a spin on the magnetic stir plate every 2 days. It took a week and a half for the jar to fully colonize.

In the Pink Buffalo jar, I couldn't see anything. I didn't see anything in the jar at all for a week. I was just about to consider tossing it when, on day 9, I saw a tiny speck floating near the bottom of the jar. Within 3 days it had substantially grown. I started stirring it every 2 days. within a week, the jar was fully colonized.

I wanted to test it on agar before reporting these results. I took a 10ml syringe full of LC from each jar. I then inoculated two plates each with .5ml. I'm pleased to report that they are all growing some nice looking mycelium!


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OfflineJoseSixpak
Djembe woof
Male


Registered: 08/19/21
Posts: 8
Last seen: 5 days, 6 hours
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: Biskop]
    #27502396 - 10/12/21 09:25 PM (1 month, 26 days ago)

I'm not familiar with your agar broth, but all my Lc's took. I did Poke tek for  all my agar to Lc's. New syringes for each one. Each PC'd in my 20 minute LC agar run. Poke for agar. Poke for Lc's Poke for clones too. my scalpel is so lonely.

Poke worked with my LMEA. PDMA, LMA+YE.

My Lc's are 400 ml water to a tablespoon of karo honey blend. (the honey bear shaped 'blend' you find at the dollar store has always worked for years.)

Keep trying. I use poke tek for everything. It's literally fire and forget. It was a game changer for me.

Good luck.


--------------------
Live by the fox. Die by the fox.


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OfflineBiskop
Fokken Prawns
Male


Registered: 12/09/17
Posts: 192
Loc: Vietnam 🇻🇳
Last seen: 6 days, 14 hours
Re: LC - No Growth After 9 Days (Josex’s Poke) [Re: JoseSixpak]
    #27503861 - 10/14/21 07:10 AM (1 month, 24 days ago)

Looking good Allen, you’re way ahead of me. I’m on my fourth batch of LC jars and haven’t seen any growth so far haha :facepalm:. I figured out that I was making it too sweet because I didn’t have anything to measure at the time. This batch I’m just using sugar water (2.8%). All the honey jars had nothing growing but still smelled okay when I opened them.

I often notice water droplets coming out of the syringe after I poke. Have you guys had anything similar happen? It could be that my biopsy just slipped out every time before it got to the jar. Just one time I felt that the needle was properly stuffed and had to use some force to get it out. I’ll be watching that jar, it’s only been 3 days I think.

Good to see you guys doing well! Been a bit busy so haven’t had time to post anything.


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