Two questions regarding media and nutrient sol'ns.

[Mr.^2]

Hey all,
I hope someone is experienced enough to help me, otherwise I have experimentation to do.

1. For gelling media agar is usually used for the substrate with liquid media nutrients. I also hear that perlite and filter paper can also be used with germination. My main question however is: has anyone used Sodium polyacrylate? If so is it recommended? I'm somewhat wary that the sodium would negatively affect the seedlings, so would a buffered sol'n help get rid of that cation?

2. Making liquid media looks like it is best made in amounts on the order of liters, amounts in excess of what I would use in a short period of time. In order the control/avoid contamination I was thinking of freezing aliquots and using as needed without drawing from the main stock. There should be no danger to freezing the nutrient sol'n, but I was wondering if anyone has tried this or if there are any better ways of controlling sol'n contamination.

I'm trying to get as much info as possible before going down the wrong path.

[Elanora]

I can't answer your first question, but as a microbiologist I can definitely address the second!

Never prepare more medium than you intend to sterilize. Growth medium is highly nutritive & very permissive to growth. Always autoclave or pressure cook medium as soon as you hydrate it. Aliquot the solution you make before sterilization (remember to leave glass jars open a quarter turn so they don't break). If you sterilize each aliquot it will be fine if you leave it at room temp as long as the container is air tight. Any growth in an air-tight aliquot indicates that your sterilization time was not long enough.

Sterilization time required depends on the volume. I never autoclave anything for less than 30 minutes. But if I have a volume over 500mL I use a 40-minute cycle, and for 1L I sterilize an hour. If you sterilize for less time all it does is allow spores to germinate, it won't actually kill them - you'll have fuzzy bottles in 2 days!

We never freeze agar solutions. I recommend aliquoting all prepared media into single use portions. If you don't have enough containers to hold a liter worth of aliquots, don't make a liter. If the metric/US measurements make it hard to calculate - 1 US cup is ~1/4 of a liter.

[Mr.^2]

Thanks for replying. I am a molecular biologist, so I have experience in autoclaving and media prep. I was just wondering if there were any issues or concerns about freezing the liquid media for supplementation. Which from what it sounds like, there shouldn't be.

Thanks again

[Elanora]

Originally Posted by Mr.^2

Thanks for replying. I am a molecular biologist, so I have experience in autoclaving and media prep. I was just wondering if there were any issues or concerns about freezing the liquid media for supplementation. Which from what it sounds like, there shouldn't be.

Thanks again

I had thought you were a biologist, but I gave the TMI version just in case you didn't do a lot of culture. I actually looked into freezing growth medium. Apparently your aliquots won't look very pretty when you pull them out of the freezer. They'll probably be a little grainy and the water may separate, so you might have to nuke 'em to get everything back in solution. But, growth shouldn't be affected. Go for it!

[Ray]

As a materials engineer - a long way from biology, no doubt - I would think the mechanics of using the acrylate gels would work against you, even if you can overcome the chemistry issues. Just guessing here, but...

In the case of liquid media, the perlite or filter paper function to keep the seeds' "heads out of the water" while providing access to the wicked-up nutrient liquid. Agar does pretty much the same thing, forming an open double-helical structure that allows the solution to sit within it. All mechanical, so far.

With the polyacrylate however, the water molecules to weakly bound to the ions in the polymer, and there is an equilibrium established with the external moisture content that controls the release. In the relatively static nature of that in the flask, it seems to me it would suck it up and never release it again, or at least somehow affect the availability to the seeds.

It seems to me it might be worth a flask or two as an experiment (assuming the polymer is OK with the sterilization temperature), but I wouldn't "bet the ranch" on it.

[Mr.^2]

Experimentation time! Thanks guys!