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pillbox
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just starting out
#23830642 - 11/13/16 09:39 PM (7 years, 2 months ago) |
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Got spores in the mail yesterday and made a homemade inoculation loop out of jewelery wire inoculated 4 dishes
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Plazmotech
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Re: just starting out [Re: pillbox]
#23830866 - 11/13/16 11:22 PM (7 years, 2 months ago) |
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Dope, good luck! For the record it looks like you have a lot of spores on that dish, probably more than necessary. No need to waste so much. Feel free to post here if you need any help.
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
Edited by Plazmotech (11/13/16 11:22 PM)
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pillbox
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That sucked. Bacteria after 48 hours.
Going to make a syrgine from the print. Any advice before I do?
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A.RichardTrickle
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Re: just starting out [Re: pillbox]
#23832591 - 11/14/16 03:07 PM (7 years, 2 months ago) |
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I'd stick with agar.... if you simply must go syringe just use the search and look up "making syringe" check off trusted cultivators and look newer than 5 years. Good luck Dick
-------------------- "When eating shit, it is best not to nibble. Bite, Chew. Swallow. Repeat." "If you're making love to your old lady, someone else is fucking her" "Douchebags are children who never grew up, like Sheeklette, we should pity them." [quote]Niffla said: [quote]A.RichardTrickle said: Dick[/quote] http://www.youtube.com/v/kbwNUOUy-3c[/quote]
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numnum59
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You could go print syringe agar and clean it up from there
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Plazmotech
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I've inoculated a few agar dishes with spore syringes too. To make a syringe it's pretty easy.
Follow the Sterile syringe tek. Put the sterile syringe after its cooled down into your still air box (SAB). An SAB is just a box with holes in it for arms. Put a shot glass and your print in there, too. Spray the inside with Lysol, and let sit for 5-10 minutes. Put on some latex gloves, alcohol them. Alcohol the outside of your syringe, and wipe down the entire shot glass with alcohol. Flame sterilize the needle. Scrape some spores from the print into the shot glass using a scalpel or whatever (make sure to flame sterilize). Spray the water from the syringe into it, then aspirate. Spray out again, and do this a few times to break up the spores. Finally aspirate it again and cap.
If you need more detailed instructions I can provide them later today. I'm on mobile right now, so it's irritating to type too much.
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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pillbox
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Re: just starting out [Re: numnum59]
#23832620 - 11/14/16 03:16 PM (7 years, 2 months ago) |
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The bacteria was just around the place I used the loop on. I Presser cooked the loop I used and flame sterilized before using. I cooled it in the agar before scrapping off some spores. Treating the spores with alcohol a good idea? Syringe to agar is the plan
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bodhisatta 
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Re: just starting out [Re: pillbox]
#23832639 - 11/14/16 03:22 PM (7 years, 2 months ago) |
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Quote:
Treating the spores with alcohol a good idea?
no
click the link in my signature, then click the agar link, then click the first one there.
you can find my pictures of how I put spores on to agar.
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Mycolorado
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Re: just starting out [Re: pillbox]
#23832661 - 11/14/16 03:29 PM (7 years, 2 months ago) |
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Quote:
pillbox said: The bacteria was just around the place I used the loop on. I Presser cooked the loop I used and flame sterilized before using. I cooled it in the agar before scrapping off some spores. Treating the spores with alcohol a good idea? Syringe to agar is the plan
You don't need to PC a loop, just flame sterilize. Anyway, if you're having issues with bacteria going from print to plate, you're going to have major bacteria/contamination issues making a syringe and then going to plate. The whole idea is just silly to me...why introduce more vectors for contamination? You might try to post up a decent pic of your plate with contamination...it might not be totally toast...
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pillbox
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Delete
Edited by pillbox (11/14/16 03:47 PM)
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pillbox
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Plazmotech
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Re: just starting out [Re: pillbox]
#23832730 - 11/14/16 03:51 PM (7 years, 2 months ago) |
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Quote:
pillbox said: Treating the spores with alcohol a good idea?
No this kills the spore.
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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pillbox
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4 dishes and no spores. None of those have bacteria..
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Plazmotech
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Re: just starting out [Re: pillbox]
#23832756 - 11/14/16 04:00 PM (7 years, 2 months ago) |
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Sorry, what do you mean?
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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pillbox
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I have 4 control dishes to make sure it was not my agar prep that failed.
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Leftfield420
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Re: just starting out [Re: pillbox]
#23832778 - 11/14/16 04:08 PM (7 years, 2 months ago) |
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At first it was hard for me to get my sterile technique down...you gotta make sure your hand is not above agar when working..keep at it, you'll be much happier with results and save materials in the long run...
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cronicr



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Knoxj up more plates to start
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
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Leftfield420
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Re: just starting out [Re: cronicr]
#23832806 - 11/14/16 04:16 PM (7 years, 2 months ago) |
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I agree with Cronicr inoplates more plates, you'll get results eventually....
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pillbox
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Maybe. Depends how far this print will last before it becomes useless
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Mycolorado
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Re: just starting out [Re: pillbox]
#23832936 - 11/14/16 04:55 PM (7 years, 2 months ago) |
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Yikes...definitely looks like it came in with the spores. Mind describing what exactly you're doing and where the print came from? Would appear to be super dirty.
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pillbox
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Got the print from a friend in another state.
A friend owns a flow hood for his farm. He let me use it for my spores. So was working in front of a hood. Ill give it another go with a syrgine the next time he has time for me to use his hood again.
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Leftfield420
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One print can do a shit load of plates....a little goes a long way
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pillbox
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Yes but if the print itself is dirty?
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A.RichardTrickle
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Re: just starting out [Re: pillbox]
#23833084 - 11/14/16 05:40 PM (7 years, 2 months ago) |
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You can clean it up on agar. You should do some reading, it would help. Dick
-------------------- "When eating shit, it is best not to nibble. Bite, Chew. Swallow. Repeat." "If you're making love to your old lady, someone else is fucking her" "Douchebags are children who never grew up, like Sheeklette, we should pity them." [quote]Niffla said: [quote]A.RichardTrickle said: Dick[/quote] http://www.youtube.com/v/kbwNUOUy-3c[/quote]
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pillbox
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Quote:
A.RichardTrickle said: You can clean it up on agar. You should do some reading, it would help. Dick
lol
Resorting to name calling. Remember barriers tell me to memeber back when I was in preK.
My question is a soild question. If the print is bacterial and I have people telling me not to make a syringe and people having me to make one. I all ready know how to make a sterile syringe and water for it.
The question is for the print.
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Leftfield420
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Re: just starting out [Re: pillbox]
#23833232 - 11/14/16 06:18 PM (7 years, 2 months ago) |
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Most prints are dirty as fuck....that's why we use agar
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pillbox
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I am giving my homemade inoculation loop an isopropyl bath right now inside a tall glass with a lid on it.
I got a shotglass, and old syringe. Going to boil the shotglass, take it out wrap in tinfoil and boil again. Use syringe from my farming buddy. I'll need to clean out one of his LC syringes. Going to suck up some boiling water a few times and empty the syringe 4+ times than boil. After that bring it to his flow hood while still in hot water. Wash shotglass with iso, and wash syringe with iso.
Take a few min break for the water to cool. Come back flame sterilize the inoculation loop. Scrape some spores place the loop into the shotglass. Flame sterilize the needle let it cool for a min. Shoot the loop with water to wash off the spores. Agitate the water and than noc up a few petri dishes.
Everything sounds right to me.
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Leftfield420
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Re: just starting out [Re: pillbox]
#23833405 - 11/14/16 07:00 PM (7 years, 2 months ago) |
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Flame it...put a drop of iso on a plate and watch the shit start growing....flaming it is best
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pillbox
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Do I use iso on the plate or do I not use iso on the plate.
People say it kills spores and not to use it.
People say use it.
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numnum59
Pro-Am Mycologist


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Re: just starting out [Re: pillbox]
#23833455 - 11/14/16 07:16 PM (7 years, 2 months ago) |
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I had a spore syringe that repetdly produced trich, eventually i was able to isolate clean myc. Probably 5-6 plates later
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numnum59
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Re: just starting out [Re: pillbox]
#23833465 - 11/14/16 07:17 PM (7 years, 2 months ago) |
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Quote:
pillbox said:
Quote:
A.RichardTrickle said: You can clean it up on agar. You should do some reading, it would help. Dick
lol
Resorting to name calling. Remember barriers tell me to memeber back when I was in preK.
My question is a soild question. If the print is bacterial and I have people telling me not to make a syringe and people having me to make one. I all ready know how to make a sterile syringe and water for it.
The question is for the print.
His name is richard, which is often shortened to dick
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Leftfield420
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Re: just starting out [Re: pillbox]
#23833494 - 11/14/16 07:27 PM (7 years, 2 months ago) |
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I was just telling you not to wipe your loop with iso....if you put a drop of iso on a plate it will contaminate.....flame your loop dude
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Leftfield420
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Re: just starting out [Re: pillbox]
#23833508 - 11/14/16 07:31 PM (7 years, 2 months ago) |
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If you are wiping your loop with iso I bet that's part of your problem there... I suggest a butane torch lighter....comes in handy when sterilizing needles and such
Edited by Leftfield420 (11/14/16 07:48 PM)
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pillbox
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It was sprayed than heat treated with a propane torch. Quote:
Leftfield420 said: Flame it...put a drop of iso on a plate and watch the shit start growing....flaming it is best
That you said put a very small amount on the plate. Made me think of getting some antibotics from a fish store to mix in while agar is cooling.
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Mycolorado
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Re: just starting out [Re: pillbox]
#23833547 - 11/14/16 07:40 PM (7 years, 2 months ago) |
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Quote:
pillbox said: I am giving my homemade inoculation loop an isopropyl bath right now inside a tall glass with a lid on it.
I got a shotglass, and old syringe. Going to boil the shotglass, take it out wrap in tinfoil and boil again. Use syringe from my farming buddy. I'll need to clean out one of his LC syringes. Going to suck up some boiling water a few times and empty the syringe 4+ times than boil. After that bring it to his flow hood while still in hot water. Wash shotglass with iso, and wash syringe with iso.
Take a few min break for the water to cool. Come back flame sterilize the inoculation loop. Scrape some spores place the loop into the shotglass. Flame sterilize the needle let it cool for a min. Shoot the loop with water to wash off the spores. Agitate the water and than noc up a few petri dishes.
Everything sounds right to me.
This is exactly why I said you'd have serious problems if you made a syringe. On top of that, don't you think if the print is dirty, it'll just make a dirty syringe? Please explain to me the reasoning behind making a syringe to go back to agar? i bet lefty is correct in his guess on the vector. You need to take the alcohol out of the equation.
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pillbox
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Well everyone is telling me to make a syringe.
So what do you recommend colo?
That is why I was thinking adding some fish antibiotics while the agar is cooling.
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Plazmotech
we love the dead



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Re: just starting out [Re: pillbox]
#23833574 - 11/14/16 07:47 PM (7 years, 2 months ago) |
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Quote:
pillbox said:
Quote:
A.RichardTrickle said: You can clean it up on agar. You should do some reading, it would help. Dick
lol
Resorting to name calling.
hahahahaha I think that's that guy's signature, he always says that at the end of each post
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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Plazmotech
we love the dead



Registered: 10/11/16
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Re: just starting out [Re: pillbox]
#23833588 - 11/14/16 07:52 PM (7 years, 2 months ago) |
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Quote:
pillbox said: Well everyone is telling me to make a syringe.
So what do you recommend colo?
That is why I was thinking adding some fish antibiotics while the agar is cooling.
I don't think you need antibiotics. I like making syringes from my spores because personally I find it easier to inoculate plates with a syringe rather than a loop.
Softer agar helps spores germinate quicker, which is a plus. The syringe will also hydrate the spores before, which might make it germinate quicker too. If you get bacteria, and its minor, don't throw away your plate just yet. Wait for the spores to germinate so you can try to get a clean (or relatively clean) wedge. That wedge will grow very rapidly onto that other plate, and in this way you can eventually isolate a clean wedge, and use it to noc a jar.
You can make a syringe if you're absolutely confident you can do so cleanly. If you think you might get even more contams by doing so, avoid it.
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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A.RichardTrickle
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Re: just starting out [Re: pillbox]
#23833599 - 11/14/16 07:55 PM (7 years, 2 months ago) |
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Quote:
pillbox said:
Quote:
A.RichardTrickle said: You can clean it up on agar. You should do some reading, it would help. Dick
lol
Resorting to name calling. Remember barriers tell me to memeber back when I was in preK.
My question is a soild question. If the print is bacterial and I have people telling me not to make a syringe and people having me to make one. I all ready know how to make a sterile syringe and water for it.
The question is for the print.
My name, is Dick... It's a play on words son
-------------------- "When eating shit, it is best not to nibble. Bite, Chew. Swallow. Repeat." "If you're making love to your old lady, someone else is fucking her" "Douchebags are children who never grew up, like Sheeklette, we should pity them." [quote]Niffla said: [quote]A.RichardTrickle said: Dick[/quote] http://www.youtube.com/v/kbwNUOUy-3c[/quote]
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Plazmotech
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I dont think you should boil the shotglass in water. Try steaming it with tinfoil wrapped around it. If you do that for even 20 minutes at 15 PSI you shouldn't need ISO. Only use ISO if you're too lazy to steam it (which has worked for me, though not great if your print is already really dirty).
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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pillbox
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Well I was thinking antibiotics to slow the growth of bacteria to allow the growth of spores than transfer away from bacteria.
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Mycolorado
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Torch loop til red hot. Cool by dipping in agar of plate. Rub loop over print. Go back to plate and swipe in a zig zag pattern on agar. You might not even see the spores that were just transferred...should have germination in around 7 days. The amount of bacteria growing on your plates where the spores were deposited and the description of your techniques lead me to believe you are contaminating the plates and not the spores. Wiping your loop and then transferring is most likely the cause if that is indeed what you were doing. If not, it's probably some other area of your transfer technique.
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numnum59
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Re: just starting out [Re: pillbox]
#23833629 - 11/14/16 08:04 PM (7 years, 2 months ago) |
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I find syringes easier to noc up agar plates than spores themselves. As for vectors, i feel your trading the noc loop for water
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pillbox
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Re: just starting out [Re: numnum59]
#23833647 - 11/14/16 08:08 PM (7 years, 2 months ago) |
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I did flame sterlize the loop between dishes. I also cooled the loop by inserting into the agar.
I might need to get a thinner loop. Mine is a little thick.
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Leftfield420
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I almost suggested the exact same thing about 10 minutes ago!
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Mycolorado
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Re: just starting out [Re: numnum59]
#23833688 - 11/14/16 08:18 PM (7 years, 2 months ago) |
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If you're just looking to dilute the spores I could see putting them in water in order to streak, but a syringe isn't necessary...and it's definitely easy to muck things up especially when you might be starting out and don't have good technique. You might try both ways but either way, you should practice your technique and once you torch an instrument it's sterile until it touches something, or something touches it. Just my 2. I hope you get some cleaner results; good luck!
Edited by Mycolorado (11/14/16 08:19 PM)
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cronicr



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Re: just starting out [Re: pillbox]
#23833695 - 11/14/16 08:20 PM (7 years, 2 months ago) |
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Quote:
pillbox said: Well I was thinking antibiotics to slow the growth of bacteria to allow the growth of spores than transfer away from bacteria.
Just make sure to eventually transfer to plain agar.
--------------------
  It doesn't matter what i think of you...all that matters is clean spawn I'm tired do me a favor
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pillbox
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http://www.ebay.com/itm/API-T-C-Tetracycline-Powder-Treat-bacterial-fish-disease-10pk-Freshwater-/252568885087?hash=item3ace47575f:g:TPIAAOSwoi1X8xtY
stable between 110-140C but really think it was me inront of the flow hood? I flame sterilized each time before I went to the spores. I made sure it was cool when I touched the agar. I got that nice sizzle sound.
I plan on only doing the spores on the anti bacteria agar than trannsfer myc to untreated agar.
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pillbox
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Re: just starting out [Re: pillbox]
#23834278 - 11/15/16 12:27 AM (7 years, 2 months ago) |
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would this or a sterilize qtip work better for applying the spores
Edited by pillbox (11/15/16 01:00 AM)
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Leftfield420
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Re: just starting out [Re: pillbox]
#23834582 - 11/15/16 07:05 AM (7 years, 2 months ago) |
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Either or...both should work....I use old tattoo needles for my loops..your loop should be fine..
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Plazmotech
we love the dead



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Re: just starting out [Re: pillbox]
#23834633 - 11/15/16 07:48 AM (7 years, 2 months ago) |
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Quote:
pillbox said:

would this or a sterilize qtip work better for applying the spores
I'm fairly certain it should be just fine. But I've heard that inoculation loops should hold the surface tension of water. In other words, they should be small enough so when you dip them in water, a thin film of water should go across the loop.
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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numnum59
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I made mine from one of those pipe cleaner things for kids art. Just burned all the frilly shit off it and whabam
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Plazmotech
we love the dead



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Re: just starting out [Re: numnum59]
#23835086 - 11/15/16 10:55 AM (7 years, 2 months ago) |
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HAHAHAHA creative
-------------------- When a man lies, he murders some part of the world. These are the pale deaths which men miscall there lives. All this I cannot bear to witness any longer. Cannot the kingdom of salvation take me home? Plazmotech's Kratom Grow
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A.RichardTrickle
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Re: just starting out [Re: numnum59]
#23835421 - 11/15/16 12:54 PM (7 years, 2 months ago) |
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Quote:
numnum59 said: I made mine from one of those pipe cleaner things for kids art. Just burned all the frilly shit off it and whabam
I'm going to try this.
-------------------- "When eating shit, it is best not to nibble. Bite, Chew. Swallow. Repeat." "If you're making love to your old lady, someone else is fucking her" "Douchebags are children who never grew up, like Sheeklette, we should pity them." [quote]Niffla said: [quote]A.RichardTrickle said: Dick[/quote] http://www.youtube.com/v/kbwNUOUy-3c[/quote]
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Mycolorado
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Registered: 07/23/16
Posts: 8,529
Loc: Interdimensional Bootcamp
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A paper clip will work...all you need is something you can flame sterilize (read: metal wire). I used some kanthal and twisted it up with a drill and allen wrench. I would like a smaller loop on mine but it works. Really though, a wire with a bent or looped tip so you don't tear the agar is all you need.
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pillbox
Stranger Danger

Registered: 11/13/16
Posts: 33
Last seen: 6 years, 5 months
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 does this look right
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pillbox
Stranger Danger

Registered: 11/13/16
Posts: 33
Last seen: 6 years, 5 months
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Re: just starting out [Re: pillbox]
#23851759 - 11/20/16 03:02 PM (7 years, 2 months ago) |
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transferred from a contaminated dish.
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Mycolorado
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Registered: 07/23/16
Posts: 8,529
Loc: Interdimensional Bootcamp
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Re: just starting out [Re: pillbox]
#23852150 - 11/20/16 05:13 PM (7 years, 2 months ago) |
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Let it grow out some and take another transfer...still looks funky and might take a few. It's best practice to put transfers in the center of the plate. Good luck!
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