You use sucrose @ 1%. I was under the ompression that Paphs did not use sucrose well, and used Fructose much better. Any thoughts? Asn what about D-inositol? Do you have any thoughts on BAP?
Yes, but a fair part of the sucrose is dissociated during autoclaving according to some chemists... I tried actually sucrose vs fructose/glucose, and sucrose made a better germination. Plus, autoclaving pure fructose is quite risky depending on what is inside the media, its reaction products can be phytotoxic...
See this:
http://www.jstor.org/pss/2482086
BAP I never use it for anything at all, there are always better, safer ways. BAP produced plants have a very high rate of mutation, and furthermore the BAP takes ages to be evacuated from the tissues once it enters. Most high standard labs avoid the use of BAP nowadays... Myo Inositol, I used to use it, but I did not find any real difference.
One more very important thing, I always make the media from scratch myself, I never buy salts preparations etc...
The MS etc... sold as a powder are not the same as the formulation, or the result of making the MS as an example by dissolving the salts 1 by 1.
As a stupid example, mix calcium nitrate and FeEDTA as powders, let the powders stand for an hour, then dilute it. After a day, you get a precipitate. If you dissolve the Calcium nitrate on one side, the FeEDTA on the other side ( never forget to boil it to dissolve it and bind it...), and add them together, it's stable. There were a few studies showing that media preparated from concentrates had a subtantial amount of precipitated things.
The second reason, the manufacturers make huge batches, and split them. As they are salts of various size, densities, etc... even if you homogeneize it for hours, it is very difficult to guarantee that a small part of the bulk will have everything in the same proportions as the bulk. That's why the suppliers provide analysis for this batch, or general formulations, but that's quite clear that, let's say sodium molybdate will not be in every 'ready to make 1L' jar sold, because the quantity for 1L will be one crystal or two... You get the same with the fertilizers too, 1g of fertilizers very, very rarely contains the proportions indicated on the bag. 1kg, most likely, 10kg, pretty sure, 25kg, definitely you should be close to the formulation.
Anyway, for the media, there are a lot of variations, how long is it sterilized, what is its composition to start with, how it is prepared, what kind of agar or gelrite, quantity per flask... All of this makes variable media ( hence I use only a few types of flasks, about 5 different ones, same for the plugs/caps, because indeed different flasks give different results and different media composition once they are properly autoclaved).
I expect too that the microwave sterilization of the media, which becomes affordable and popular, will make even more problems, because our formulations were made before with an autoclaving of 10-25min and 115-132 degrees celsius, which definitely made the final product different from the prepared media. With those new technologies like microwave or microfiltration, the final results are way different...
. Perhaps someone could tell me if they (species) are difficult or not.
