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OfflinePippen
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Registered: 12/17/19
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Last seen: 3 years, 10 months
Struggling with agar (first timer)
    #26432069 - 01/13/20 08:02 PM (4 years, 1 month ago)

First timer here and my agar isn’t looking like I expected. I inoculated on 1/5 and took these pics just now. It was really hard to even get anything to show up in the pics. Basically all I can see is a very faint change in texture on the surface. There’s also some red discoloration following the path that I dragged the inoculation loop through. I was expecting it to look cloudier and white.

I used 6g brown rice flour, 9g agar, and 400mL water. PC at 15 psi for 25 mins (10 min venting). Inoculated in a SAB and tried my best to observe clean techniques but it was my first time so I’m sure I could have done better. I dipped the loop in 70% alcohol and then torched it. Cooled it in agar then dipped in the solution from the spore syringe and made a “z” pattern in the agar. The dishes were wrapped and have been sitting in a 70 F room with 12 hours of fluorescent light per day.

Any help please?



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OfflineJarhead3521S
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Re: Struggling with agar (first timer) [Re: Pippen]
    #26432087 - 01/13/20 08:11 PM (4 years, 1 month ago)

I’ve usually used 9g BRF and 9g agar, roughly 2%.....seems 6g would be on the less nutritious side but maybe somebody with more knowledge will step in.  I recently switched to lme but used same percentage.


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OfflineMLPismyOPSEC
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Re: Struggling with agar (first timer) [Re: Pippen]
    #26432089 - 01/13/20 08:12 PM (4 years, 1 month ago)

Could you elaborate on your method when you say you dipped the loop in the solution? Did you squirt the syringe into another container or something? IMO, being from spore solution, it's too early to tell. It usually takes me two weeks to see anything substantial.


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OfflinePippen
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Re: Struggling with agar (first timer) [Re: Jarhead3521]
    #26432101 - 01/13/20 08:18 PM (4 years, 1 month ago)

Quote:

Jarhead3521 said:
I’ve usually used 9g BRF and 9g agar, roughly 2%.....seems 6g would be on the less nutritious side but maybe somebody with more knowledge will step in.  I recently switched to lme but used same percentage.




I had found someone that recommended this ratio but maybe it’s not optimal. I was thinking of trying light malt extract next.

Quote:

MLPismyOPSEC said:
Could you elaborate on your method when you say you dipped the loop in the solution? Did you squirt the syringe into another container or something? IMO, being from spore solution, it's too early to tell. It usually takes me two weeks to see anything substantial.




Yes, I squirt some into another petri and then dip the loop in there. Maybe I’m being too impatient. I saw people saying to expect growth within 4 days but others that said it could take weeks so I didn’t really know what to expect. It seems like I have really weak growth so I thought maybe something was wrong.


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OfflineHighHarles
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Re: Struggling with agar (first timer) [Re: Pippen]
    #26432105 - 01/13/20 08:20 PM (4 years, 1 month ago)

First thing this hobby teaches you patience, I wish you luck!:rasta:


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OfflinePippen
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Re: Struggling with agar (first timer) [Re: HighHarles]
    #26432115 - 01/13/20 08:23 PM (4 years, 1 month ago)

Quote:

HighHarles said:
First thing this hobby teaches you patience, I wish you luck!:rasta:




Thank you :smile: Patience is not my strongest trait haha. I’ve been staring at these things way too much. I decided to inoculate 12 more dishes with other syringes the other day to see if they fair any better. Then I made 25 more agar dishes. I’m ready to put the work in. Just want these things to take off :wink:


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OfflineJarhead3521S
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Re: Struggling with agar (first timer) [Re: Pippen]
    #26432129 - 01/13/20 08:29 PM (4 years, 1 month ago)

The BRF is fine, I would just add a few more grams.  I’ve gone through about 80 plates with that ratio and have had great growth, MS is slow though for sure.


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OfflineMLPismyOPSEC
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Re: Struggling with agar (first timer) [Re: Pippen]
    #26432136 - 01/13/20 08:34 PM (4 years, 1 month ago)

Quote:

Pippen said:
Yes, I squirt some into another petri and then dip the loop in there.




This could also be why, by squirting the solution into the empty petri and dipping the loop, it might not be picking up enough spores. Theoretically, it shouldn't be a problem, but you'd have a little better luck by using the syringe to drop one drop on the blank petri and smear it with the loop. But yeah, definitely need patience! You will learn, but it will be a hard habit to break, i still struggle with it a year in :lol:


Edited by MLPismyOPSEC (01/13/20 08:36 PM)


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OfflinePippen
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Re: Struggling with agar (first timer) [Re: MLPismyOPSEC]
    #26432225 - 01/13/20 09:22 PM (4 years, 1 month ago)

Quote:

Jarhead3521 said:
The BRF is fine, I would just add a few more grams.  I’ve gone through about 80 plates with that ratio and have had great growth, MS is slow though for sure.




Ok thank you I’ll bump it up next time.

Quote:

MLPismyOPSEC said:
Quote:

Pippen said:
Yes, I squirt some into another petri and then dip the loop in there.




This could also be why, by squirting the solution into the empty petri and dipping the loop, it might not be picking up enough spores. Theoretically, it shouldn't be a problem, but you'd have a little better luck by using the syringe to drop one drop on the blank petri and smear it with the loop. But yeah, definitely need patience! You will learn, but it will be a hard habit to break, i still struggle with it a year in :lol:




Thanks for the tip!


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