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OfflineRichard_D_James
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Registered: 04/27/01
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How to mix Peroxidated MEA
    #582784 - 03/18/02 11:23 PM (14 years, 8 months ago)

WHat is the best way to mix up Peroxide in cooled MEA?  I can't seem to get even distribution... what do the pros use?

:smile:

thx in advance


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OfflineAzure
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Re: How to mix Peroxidated MEA [Re: Richard_D_James]
    #582878 - 03/19/02 01:31 AM (14 years, 8 months ago)

I don't know what pros do, but here's what I do:
First, I mix up agar, malt, potato dextrose and melt the solution on the stove. I then place the mixture in a 500 ml flask, seal the top with a double layer of tinfoil, and pressure cook at 15 psi for 10 minutes. I allow the cooker to release all the pressure, and when the mixture is still hot, I take it out of the cooker. When the flask is cool enough to touch without burning your fingers, I add 7% H2O2(which is a 3% concentration H2O2 when it comes in the bottle) to the liquid malt and swirl the flask until the peroxide is evenly distributed.
If I use my pyrex petri dishes, I sterilize the clean, empty dishes with the flask. If I use plastic petri dishes, I merely open the package and use them immediately because they come sterile. I pour the dishes on an anal retentively sterilized surface(or at least pretty close to sterilized) and allow them to solidify. Once they are solid, I use the oven and transfer sterile tissue to the new plates, and then seal the plates with parafilm. Saran wrap works also if you don't have such luxuries.
When using 10%, I noticed the tissue samples did not like it too much and stalled for weeks. With a 7% concentration, contams can still occur if you're not careful, but at least the majority are killed and the tissue adapts rapidly to the medium.


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OfflineAnnoA
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Re: How to mix Peroxidated MEA [Re: Azure]
    #582898 - 03/19/02 01:58 AM (14 years, 8 months ago)

7% concentration? and this works??

Wayne says to use no more than 5 ml of the 3% peroxide per 500 ml of agar.


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OfflineMAIA
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Re: How to mix Peroxidated MEA [Re: Richard_D_James]
    #583080 - 03/19/02 07:43 AM (14 years, 8 months ago)

There was a thread in this forum about this SAME issue this last week. Search for it....

Peace,
MAIA


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Offlinekarlfinn
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Re: How to mix Peroxidated MEA [Re: Azure]
    #583086 - 03/19/02 08:11 AM (14 years, 8 months ago)

right....rush says 6-8ml / liter water, broth whatever...so 3.5 ml / 500 ml agar mix...is your .7 %///
used this in my liquid culture and mycelia floated to top where it was easy to
syringe out through the hole.......but now the remaining mycelia culture, 24 hrs later,
has matted up ....pretty solid...expect some recovery time, but....
any experience with this.??..I/l post as a question...later...I guess...farmer


Edited by karlfinn (03/19/02 05:07 PM)


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OfflineAzure
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Re: How to mix Peroxidated MEA [Re: Anno]
    #583443 - 03/19/02 04:36 PM (14 years, 8 months ago)

It definitely works, but the reason it does is probably because the solution is still relatively hot when I add the peroxide to it, which breaks down the the peroxide and lowers the molarity.


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Offlinekarlfinn
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Re: How to mix Peroxidated MEA [Re: Azure]
    #583488 - 03/19/02 05:15 PM (14 years, 8 months ago)

azure....is that 7 % or . [point] 7 % ?... one is 7 ml pox / liter , ...or...other;... 70 ml pox / liter..???
all real new to me but I just the lesser amount..??


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OfflineParticleMan
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Re: How to mix Peroxidated MEA [Re: karlfinn]
    #591085 - 03/27/02 04:43 PM (14 years, 8 months ago)

word. when i was reading earlier someone suggested the use of a candy thermometer, stick it in your agar mix whatever it may be, wait till it is at 140 deg. then pour in the H202, at this temp the agar mix is still liquid, yet cool enough not to break down the H202.


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