Microwave sterilization

Paul N Hengen pnh at ncifcrf.gov
Wed Dec 18 11:48:32 EST 1996


Michael J. Prigge (prigge at darkwing.uoregon.edu) wrote:

>>> From what I understand moisture is essential for the sterilisation
>>> process.  Another poster described a microwave based sterilisation
>>> unit that involved the generation of steam.  I don't think that
>>> simply microwaving dry glassware in the oven has any effect.
>> 
>> I don't think so either. 
> 
> I beg to differ on this point... a method I have used to sterilize
> Arabidopsis seed is essentially dry. I nuked 15 ml polyprop tube 
> with a few hundred seed twice for five min each time. At the end 
> of the dose, the tube was kinda soft but the seed was ~100% viable 
> AND NO FUNGUS (or anything else) GREW when plated.
> 
> Now for the potential caveats: I plated the seed onto kanamycin
> or hygromycin plates; and there was a beaker of water in the 
> microwave--to soak up unquenched microwaves(?)--that was replaced as 
> it began to boil. Without sterilization, though, kan and hyg plates 
> are overrun with fungii before germination.
> 
> Evidently, the water content of the bacteria and spores and lack 
> thereof in the dehydrated seed embryos contributes to the 
> differential killing. I know that, if weed contamination is a 
> problem for you, longer runs take care of the Arabidopsis too. :)

Interesting! I wonder if you have the lid of the tube on tight while the seeds
are being zapped. My point would be that the steam is getting in to the seed
coats and that's what kills your fungi (?).

Anyway, the original idea about microwaving a flask or beaker made me think
about what that person was using it for. If it is to grow a bacterial culture
in sterile broth for plasmid DNA (vector) isolation, I can admit that in the
past I have not sterilized media for this at all. I was appalled once when my
mentor added dry media components to a non-sterile flask, added water, then
immediately innoculated it with log-phase bacteria. Being trained as an
undergrad to sterile everything, I was shocked! Since then, in a pinch I'll
admit I've done the same thing. Anyone else do this?  Because the culture is
growing so rapidly, the bloom occurs within minutes and tops off within 2-3
hours. I then collect the bacteria and do my plasmid prep.

--
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* Paul N. Hengen, Ph.D.                           /--------------------------/*
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