I prepared M9 medium by autoclaving the M9 salts and glucose separately. Calcium chloride, magnesium sulfate, and trace minerals were dissolved in autoclaved water and sterilized by filtration. However, mixing these reagents together and preparing M9 media is causing the solutions to have a very light yellow color. It's been two weeks since I prepared this solution and wanted to know if this is normal. Thanks..
The M9 medium typically contains Minimum Essential Medium (MEM) Vitamin Solution (100x), which is yellow. Furthermore, a small amount of FeCl3 can be also expected to contribute to the observed colour. Please check if the mentioned media was somehow modified, namely with the addition of a pH indicator, or in some other way that may affect its colour. CaCl2 is prone to atmospheric carbonation (assisted by humidity) what should be avoided; you may want to check the pH of the freshly prepared CaCl2 salt solution (*). Contaminated CaCl2 salt could possibly result in partial calcium precipitation (possibly as CaCO3), and/or in colloidal suspended particles, with some tendency for glucose (perhaps with other biochemical components) to become chemisorbed to some extent, while the ferric ion might possibly play a coagulant role ― if its co-precipitation as hydroxide can also occur. That might affect experimental reproducibility, even considering that the buffering capacity of the media should be able to prevent any significant pH deviation.
(*) You may want to check: https://www.researchgate.net/post/whats_the_best_choice_to_modify_the_PH_of_calcium_chloride_hexahydrate https://www.researchgate.net/post/Why_is_my_anhydrous_calcium_chloride_not_going_into_solution
Hi,
We are having a similar problem with M9 in our lab. after adding the trace metals the media turns yellowish and a bit cloudy. when adding carbon sources to the media, a yellow precipitation appears after a day. Carlos's explanation seams to fit, but how can one avoid the problem? any advice will do.
Our recipe by order (1.1X M9, before adding carbon sources): 35ml, miliQ H2O
5µl, 1M calcium chloride (CaCl2)
100µl, 1M magnesium sulfate (MgSO4)
50µl, 1000x Trace metals
10ml, 5X M9 salts
Thanks a lot!
Hi Nesli. I have just decided to prepare the media fresh every time I use it. But I think the issue comes from the trace minerals. You might try mixing everything else and leaving the trace minerals out. You can just add that right before use.
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