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rxb
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rxb agar jar tek - low cost low tek isolation solution 2
#21899443 - 07/05/15 01:31 AM (8 years, 6 months ago) |
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this will allow you to do isolation without a flow hood, you dont even really need a still air box, but using one wouldnt hurt anything and might help, who knows. flow hoods are great, im not knocking flow hoods. this should give you as good of results, you can even take isolated material from a mold contaminated plate without blowing mold spores all over your lab.
for most applications i prefer this method to using a flow hood, but it isnt as easy to see what you are doing, and you dont get as many style points.
you do however save hundreds of dollars 
Things you need: 1) 8 oz widemouth jars with lids (they come 4 for $4 at walmart) 2) a 2+ inch needle, I use a 12 gauge 2 inch autoclavable stainless steel needle. (there are vet supply places which have them for $2, they are reusable so i dont really count this as a cost but shrug) 3) aluminum foil ($1) 4) agar (you wont even use a dollars worth per jar) 5) a straight pin, or small gauge needle for punching air holes. (free) 6) high temp silicone from an auto parts shop (optional, but i like it) 7) poly fill (free in a pillow, or 2 dollars at a fabric store)
i used a quart jar for the construction demo, but this should be done in 8oz widemouth jars, so they are short enough for this to work, all my 8 oz jars were full... you should get the idea 

1) mix agar.
2) fill your jars with 50 cc of mixed agar (you can find 60cc syringes at tractor supply)
3) cut an aluminum sheet the width of the jar or more and fold it in half so that its two sheets, then do this again, so you have 2 aluminum rectangles that are 2 layers each.

4) get your jar, and put one aluminum foil set on it, and create a little bowl in the foil and crimp along the edge of the jar

5) take a polyfill ball about the size of a baseball and smoosh it in your hands until its very small

6) attempt to put the now tiny polyfill ball in the bowl created, and then cover with the second foil rectangle

7) while struggling desperately to keep the polyfil ball under the second aluminum sheet put the ring on top, stuff in any polyfill that sticks out and then push down and tighten the jar ring.

8) using a straight pen (or anything that can pierce holes in the aluminum) poke lots of holes and be sure to get through both layers of foil. this is pressure relief and air exchange without holes it will probably not survive the PC

9) repeat process until all jars are done.

10) pressure cook the jars at 15 psi for 30 minutes. Allow pressure cooker to reach 0 psi before opening the pressure cooker.

11) allow ample time for jars to cool to room temperature.
12) apply spores or live culture to the agar with a syringe. If you have dry spores, mix with sterilized water, then suck into syringe and drip onto agar. Use a similar process for tissue samples, live culture and spore syringes can be used directly, be sure to flame needles.
13) after injection, if you use a large needle, its not a bad idea to put a small dot of high temp silicone over the hole created by the needle.
14) wait until the dish is colonized to your satisfaction.
15) fit a syringe with your large guage 2 inch long or greater needle (I use 12 gauge 2 inch stainless steel needles I get from an online vet supplier) and fill the syringe with sterilized water.
16) because we are using foil and a very short jar, a specific sector can be isolated by putting the needle into the jar right above the sector. Make a small circle with the needle around the sector to be sampled as a boundary layer. And scratch a grid across the top of the sector

17) be sure you are on a level work surface and put a few drops of water in the sector and stir the water into the agar then add another drop or two. Then pull the water back into the syringe and repeat the process until you are satisfied you have enough material.
18) remove from the jar, shake the syringe well, and flame sterilize the needle.
19) drip the contents into new agar jars.
20) repeat the process until you have an isolated mycelium colony.
21) once you have an isolated strain you can make a live culture by simply adding sterilized water and shaking the jar and then removing all water, sometimes the mycelium will recolonize the agar after you do this, but it usually breaks up the agar.
not the best at writing so im sure i left a few typos, thanks for reading
Edited by rxb (07/11/15 11:55 PM)
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931186 - 07/11/15 10:25 PM (8 years, 6 months ago) |
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i have examples of spores in the agar jars and a transfer from multispore syringe to multispore culture and an isolation from the culture.. the problem is i have not figured out how to photograph the jars in a way that looks like ANYTHING.
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931189 - 07/11/15 10:27 PM (8 years, 6 months ago) |
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i suppose i'll eventually just do some jars i dont care about and open them up and photograph them.
at the moment im just enjoying that this is easier than my flow hood, and seems to be working.
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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azur
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931385 - 07/11/15 11:27 PM (8 years, 6 months ago) |
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Easier than your flow hood? I missed something. I didn't think if could get easier than a flow hood. It is a promising tek though. Well thought out and performed. Good job
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: azur]
#21931391 - 07/11/15 11:28 PM (8 years, 6 months ago) |
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i cant use the flow hood on my couch.
can you?
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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azur
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931404 - 07/11/15 11:30 PM (8 years, 6 months ago) |
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Not quite;)
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931406 - 07/11/15 11:31 PM (8 years, 6 months ago) |
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i guess if i moved the flow hood beside the couch... they would be equally easy
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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azur
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931416 - 07/11/15 11:33 PM (8 years, 6 months ago) |
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Either way, though I'll never use this tek, it's great for beginners that find sab's cumbersome and don't have loot for a hood. You should have it moved to cult. Probably do better there
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: azur]
#21931439 - 07/11/15 11:38 PM (8 years, 6 months ago) |
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you might try it one day just cuz its great for long term storage as little jars of colonized agar are like huge slants
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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azur
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931450 - 07/11/15 11:40 PM (8 years, 6 months ago) |
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I may make some in the little jars
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: azur]
#21931457 - 07/11/15 11:43 PM (8 years, 6 months ago) |
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yeah the little jars are kind of the selling point... little jar + big needle = isolation without $$
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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azur
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: rxb]
#21931474 - 07/11/15 11:46 PM (8 years, 6 months ago) |
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution [Re: azur]
#21931483 - 07/11/15 11:48 PM (8 years, 6 months ago) |
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ps thanks for bumping the shit out of my tek
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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Prisoner#1
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21931500 - 07/11/15 11:50 PM (8 years, 6 months ago) |
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This thread was moved from Advanced Mycology.
Reason: moved at user request
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azur
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: Prisoner#1]
#21931514 - 07/11/15 11:52 PM (8 years, 6 months ago) |
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We're gonna be tired in the morning pris.
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azur
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: azur]
#21935266 - 07/12/15 09:44 PM (8 years, 6 months ago) |
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Bumping to share
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Mr.GuessWork
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: azur]
#21936511 - 07/13/15 09:15 AM (8 years, 6 months ago) |
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The use of the syringe to pick up the isolate is a pretty cool idea. Isn't it kind of hard to do a streak to get good separation between colonies with all that stuff in the way though?
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FreeWorldOrder


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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: Mr.GuessWork]
#21936674 - 07/13/15 10:14 AM (8 years, 6 months ago) |
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While this could work okay for transfers & myc. for inoculations, it seems it may be tough to isolate sectors like this... 
SAB still seems much easier than going through all the process of making up the jars and trying to isolate with a syringe needle through the 2 layers of foil and polyfil.
Not saying it can't work, just seems like a lot of effort & time when one can just use a $7 SAB and do the same thing in just minutes or even seconds...
-------------------- "They who can give up essential liberty, to obtain a little temporary safety, deserve neither liberty nor safety." - Benjamin Franklin Lets Grow Mushrooms Videos PastyWhyte's Easy Agar TEK Agar's Liquid Inoculant TEK
Edited by FreeWorldOrder (07/13/15 10:21 AM)
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: FreeWorldOrder]
#21937254 - 07/13/15 01:04 PM (8 years, 6 months ago) |
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Quote:
FreeWorldOrder said: While this could work okay for transfers & myc. for inoculations, it seems it may be tough to isolate sectors like this... 
SAB still seems much easier than going through all the process of making up the jars and trying to isolate with a syringe needle through the 2 layers of foil and polyfil.
Not saying it can't work, just seems like a lot of effort & time when one can just use a $7 SAB and do the same thing in just minutes or even seconds...
it works PERFECTLY for isolating sectors.
because you can put the needle anywhere on the lid, you can get material from very very specific locations. thats really the POINT of the tek.
i have a number of SAB and a flow hood, and a posative pressure box. putting the needle through 3 layers is effortless. isolation is effortless. WAAAAAAY easier than a SAB in my opinion.
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
Edited by rxb (07/13/15 01:06 PM)
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Mr.GuessWork
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937299 - 07/13/15 01:16 PM (8 years, 6 months ago) |
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How do you inoculate the plate with the needle? Do you only do liquid inoculation? It seems's like it'd be hard to get a good streak, especially if you wanted to do a couple streaks to get really good separation.
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: Mr.GuessWork]
#21937358 - 07/13/15 01:36 PM (8 years, 6 months ago) |
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i mix spores with water.
i drip spore syringes in multiple spots.
also it has this neat feature where if you put a quarter cc from a spore syringe on the agar... (enough to get a good bead of water)... you can then roll the bead around.... over the whole plate... then turn it upside down for a day or two (so the bead doesnt sit on the agar)... and the whole plate gets COVERED In colonies.
so your techniques just adapt to the situation.
but you know, if its not your idea of a good time, you dont need to try it 
but i think if you do, you'll be pleasantly surprised at how simple it is to do whatever you'd like
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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spacechildo
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937383 - 07/13/15 01:42 PM (8 years, 6 months ago) |
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how do you get myc out of the agar plate if you dont use a SAB?
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937386 - 07/13/15 01:42 PM (8 years, 6 months ago) |
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forgive the horrible picture
this was done thursday or friday night with that roll the bead around technique.
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
Edited by rxb (07/13/15 01:42 PM)
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: spacechildo]
#21937395 - 07/13/15 01:44 PM (8 years, 6 months ago) |
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Quote:
spacechildo said: how do you get myc out of the agar plate if you dont use a SAB?
from the tek
Quote:
16) because we are using foil and a very short jar, a specific sector can be isolated by putting the needle into the jar right above the sector. Make a small circle with the needle around the sector to be sampled as a boundary layer. And scratch a grid across the top of the sector
17) be sure you are on a level work surface and put a few drops of water in the sector and stir the water into the agar then add another drop or two. Then pull the water back into the syringe and repeat the process until you are satisfied you have enough material.
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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spacechildo
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937405 - 07/13/15 01:46 PM (8 years, 6 months ago) |
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have you transfered away from contams successfully like that?
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: spacechildo]
#21937408 - 07/13/15 01:47 PM (8 years, 6 months ago) |
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Quote:
spacechildo said: have you transfered away from contams successfully like that?
nope, i havent contaminated one yet. - i will contaminate one on purpose, i have a liquid culture with green mold, i just have to do it and have not yet
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
Edited by rxb (07/13/15 01:48 PM)
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spacechildo
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937421 - 07/13/15 01:50 PM (8 years, 6 months ago) |
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it sounds real risky to me. just cant see it being a feasable way of dealing with agar transfers..
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: spacechildo]
#21937426 - 07/13/15 01:51 PM (8 years, 6 months ago) |
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Quote:
spacechildo said: how do you get myc out of the agar plate if you dont use a SAB?
Quote:
spacechildo said: it sounds real risky to me. just cant see it being a feasable way of dealing with agar transfers..
some people have to try things before they understand how they work.
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937430 - 07/13/15 01:53 PM (8 years, 6 months ago) |
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im not sure exactly what you are risking?
the 2 drops from a spore syringe?
the 10 bux or less it costs to make?
the 10 minutes it takes to make one?
or just your illusions that you cant work without a flow hood?
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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spacechildo
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937434 - 07/13/15 01:53 PM (8 years, 6 months ago) |
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exactly!
report back once you've tried transfering away from contams
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: spacechildo]
#21937437 - 07/13/15 01:54 PM (8 years, 6 months ago) |
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I SURE WILL!
you do the same!
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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rxb
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937462 - 07/13/15 02:02 PM (8 years, 6 months ago) |
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part of the reason i havent done this yet, is that the above photo looks TERRIBLE. i have good camera equipment and i need to get it out and set things up and take GOOD photos.
so i've done a lot of peat and repeat.
but i'd like to be more show and tell.
i'll knock one up with mold this week...and i'll try to get the real camera setup.
-------------------- ->$10 FLOW HOOD ALTERNATIVE <- . i cleaned a mold contaminated live culture and saved it. (might have useful applications) [quote]Enlil said: I'd be the guy with thousands of minions doing my bidding and all of the hot women locked in a cage for my use.[/quote]
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Mr.GuessWork
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Re: rxb agar jar tek - low cost low tek isolation solution (moved) [Re: rxb]
#21937491 - 07/13/15 02:11 PM (8 years, 6 months ago) |
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Quote:
rxb said: i mix spores with water.
i drip spore syringes in multiple spots.
also it has this neat feature where if you put a quarter cc from a spore syringe on the agar... (enough to get a good bead of water)... you can then roll the bead around.... over the whole plate... then turn it upside down for a day or two (so the bead doesnt sit on the agar)... and the whole plate gets COVERED In colonies.
so your techniques just adapt to the situation.
but you know, if its not your idea of a good time, you dont need to try it 
but i think if you do, you'll be pleasantly surprised at how simple it is to do whatever you'd like 
It's definitely got some advantages. It probably doesn't waste as much of the agar as a 3 or 4 streak method does. I think I'll try something like it next time I play with agar. Thanks for the inoculation specifics.
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