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TrippyMomp



Registered: 04/16/20
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spores VS brf pf tek VS agar for intial inoc 1
#26648306 - 05/04/20 12:45 PM (4 years, 1 month ago) |
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Pretty sure my pt syringes either got irradiated in the mail or are weak af. Made a 5 brf cakes to see if my pt would work. nothing really happened on my agar with the pt. with my GT and EC both had lots of growth but allot looks like bacteria so im trying to figure out the best way to initially star or what method of best success. I have a SAB and used sterile technique. Transferred some promising growth to new agar plates one I took an agar chunk via scapel a few others I just stabbed the single colonies since the plates looked flooded with diff colonies I figured I need to isolate for one to grow stronger looked like thousands. My GT syringes seem the strongest so I put a whole syringe in another batch of BRF cakes today 10 of them in the wide mouth mason jars. Looking at the flow chart it appears spore to BRF is best because its less susceptible than other initial methods. MY main question is am I gonna have the best luck right now with BRF since im working with potentially dirty spore singes till I get going and am wasting my time on AGAR right now till I have cleaner samples to use ?
thanks guys also just for fun put an agar plate to my air vent to see what comes up lol.
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp]
#26652474 - 05/06/20 08:30 AM (4 years, 1 month ago) |
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Alll by my seeeeeeeelf......
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poisoned
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp]
#26652490 - 05/06/20 08:38 AM (4 years, 1 month ago) |
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agar is the most sure way to get clean cultures. Just got to keep transferring away from contams.
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: poisoned]
#26652557 - 05/06/20 09:18 AM (4 years, 1 month ago) |
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Quote:
poisoned said: agar is the most sure way to get clean cultures. Just got to keep transferring away from contams.
Do these look promising I transferred them and they seem to be doing more the smaller one not in the wedge was via stab inoculation I sterilized a stab tool in my dissection kit and just poked the center of one of the white dots on my old more contaminated looking plates think from the syringe water. Should I scrape the myc off this wedge or just leave it as is. Looks like a little bacteria maybe came with ? Thanks for the response man.
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp]
#26652565 - 05/06/20 09:21 AM (4 years, 1 month ago) |
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Should be getting premade plates today I know lazy lazy buy also have the stuff to make more just dong want it to go bad in the fridge. Do you think I should let these develop more before I transfer anything theres really not segments yet so it's not very large yet. Also do you think micropore tape is hindering AE or do plates not need it. Just scared to not have it taped need to get Seran wrap but keep forgetting
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poisoned
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp]
#26652568 - 05/06/20 09:22 AM (4 years, 1 month ago) |
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There seems to be bacteria on both plates. You could try doing a sandwich or hot pour. I've rescued a culture from a shit ton of contamination with a very badly done sandwich method.
sandwich method is basically covering your growth with clean agar from another plate. Myc will penetrate agar while bacteria won't.
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poisoned
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Re: spores VS brf pf tek VS agar for intial inoc [Re: poisoned] 1
#26652570 - 05/06/20 09:23 AM (4 years, 1 month ago) |
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Micropore tape works for plates. It's just a bit of a hassle
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: poisoned]
#26652575 - 05/06/20 09:27 AM (4 years, 1 month ago) |
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Like with a scalpel? Sorry probably dumb question but do I remove the clean agar from the plastic plate it's in and sandwhich it on top ? Are allot of syringes from vendors on this site like that or are they all just in general and bacteria is inevitable and must be removed via agar
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poisoned
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp]
#26652581 - 05/06/20 09:31 AM (4 years, 1 month ago) |
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https://www.shroomery.org/forums/showflat.php/Number/2823255
I did it with normal MEA agar and without peroxide and it worked. I rarely get any contams from spores, but it happens. You could also try inoculating another plate with a zigzag technique and see if you can get any growth away from bacteria.
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TrippyMomp



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Re: spores VS brf pf tek VS agar for intial inoc [Re: poisoned]
#26652654 - 05/06/20 10:14 AM (4 years, 1 month ago) |
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Yeah maybe this next syringe of gt will be less contaminated. Makes me worry for these 10 brf cakes I knocked up
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: poisoned]
#26654068 - 05/06/20 11:20 PM (4 years, 1 month ago) |
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Quote:
poisoned said: https://www.shroomery.org/forums/showflat.php/Number/2823255
I did it with normal MEA agar and without peroxide and it worked. I rarely get any contams from spores, but it happens. You could also try inoculating another plate with a zigzag technique and see if you can get any growth away from bacteria.
So I'm gonna do this tomorow my mycellium is pretty small. My plates are decently thick tonight and cooling in my pc glad twist n lock minis. 2 I made smaller but still are atleast 1/2 cm maybe 1/10 of in h thick. It should work no matter what just might take longer if its thick correct?
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp]
#26654094 - 05/06/20 11:39 PM (4 years, 1 month ago) |
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And is 3% peroxide worthless
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LogicaL Chaos
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp]
#26654110 - 05/06/20 11:52 PM (4 years, 1 month ago) |
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Could be your vendor. Some vendors make very dilute syringes. Try a different vendor.
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CocaineBuffet
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Re: spores VS brf pf tek VS agar for intial inoc [Re: LogicaL Chaos] 1
#26654165 - 05/07/20 12:36 AM (4 years, 1 month ago) |
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It would help if you shared the pics of the first plates. It looks like you may have taken transfers before mycelium had a chance to show itself based on the transfers
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: CocaineBuffet]
#26654213 - 05/07/20 01:12 AM (4 years, 1 month ago) |
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Quote:
CocaineBuffet said: It would help if you shared the pics of the first plates. It looks like you may have taken transfers before mycelium had a chance to show itself based on the transfers

I think its all pretty juvenile and needs to grow I also don't have the Mycology eye yet. That's the plate I took the transfer from I cut out the best looking spot on there and transferred it here.
 I stabbed the other one and took a tiny chunk of agar with it and now its also colonizing.. 
Heres some of the possibly contaminated older plates of mine. like I said I have 9 in my PC right now cooling and would like to try to move forward productively tomorrow just looking for advice and personal opinions. Ill put up the more promising looking ones to me some i accidently flooded first batch then second batch i learned and used a loop inoculation method and worked much better but people still said lots of bacteria. are all these spots bacteria or just he like more circular larger splotches that have less depth and wider surface area.? anyway heres some pics I have GT, PT, EC. Think my PT got irradiated in the mail. all my syringes have lots of visible spores in them. 1. 2. 3. 4. 5. 6.flooded forsure 
ALSO DOES THIS LOOK LIKE MYC ? TRYing to revive the PT in a LC first then Agar isolate.
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TrippyMomp



Registered: 04/16/20
Posts: 185
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Re: spores VS brf pf tek VS agar for intial inoc [Re: LogicaL Chaos] 1
#26654214 - 05/07/20 01:14 AM (4 years, 1 month ago) |
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Quote:
LogicaL Chaos said: Could be your vendor. Some vendors make very dilute syringes. Try a different vendor.
True maybe just bad luck. hopefully these BRF cakes take and I have a back up plan ! just trying to get some prints and what not for long-term. All the advice is much appreciated.
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mushpunx
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp] 2
#26654428 - 05/07/20 05:37 AM (4 years, 1 month ago) |
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Micropore tape really sucks for wrapping plates. It's a huge pain.
The best option is Parafilm, that's what it is made for, but it's expensive.
What is easiest to do, is take a roll of plastic cling wrap and a sharp knife, and saw a 1" section off the end of the roll. Really useful for wrapping plates.
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TrippyMomp



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Re: spores VS brf pf tek VS agar for intial inoc [Re: mushpunx] 1
#26654701 - 05/07/20 09:10 AM (4 years, 1 month ago) |
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Quote:
mushpunx said: Micropore tape really sucks for wrapping plates. It's a huge pain.
The best option is Parafilm, that's what it is made for, but it's expensive.
What is easiest to do, is take a roll of plastic cling wrap and a sharp knife, and saw a 1" section off the end of the roll. Really useful for wrapping plates. 
deff gonna get some saran wrap today. On a better note I believe my PT are actually doing something in 2 of the 5 jars day 7 knocked up. 4/30/20 knock up PT-   
Also was thinking of doing the sandwhich agar technique for the one i transferred on the wedge. the smaller one i think would be more suitable for a hot pour so im prepping agar in my pc right now any thoughts ? maybe I should just hot pour them both ? Some of my glad twist n locks/most came out with a slant on them lol i need a bigger PC but also just want smaller agar containers
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: TrippyMomp] 1
#26654705 - 05/07/20 09:12 AM (4 years, 1 month ago) |
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Also using the wide mouths for my GTs. figured id use these shit ones with the PTs cause I wasn't sure if they'd germ or not, and gray on jars is water marks from PC.
Edited by TrippyMomp (05/07/20 09:13 AM)
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TrippyMomp



Registered: 04/16/20
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Re: spores VS brf pf tek VS agar for intial inoc [Re: poisoned]
#26654752 - 05/07/20 09:44 AM (4 years, 1 month ago) |
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Quote:
poisoned said: There seems to be bacteria on both plates. You could try doing a sandwich or hot pour. I've rescued a culture from a shit ton of contamination with a very badly done sandwich method.
sandwich method is basically covering your growth with clean agar from another plate. Myc will penetrate agar while bacteria won't.
how likely is hot pour to kill the myc from the heat. thinking bout doing it on the smaller one that's not the wedge along with the plate I took the wedge from because its showing more myc but in a pool of bacteria these damn syringes. Makes me worry about my PTs in the BRF right now lol since I believe those to be the dirtiest.
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