I studied browning of the madia and encountered a problem in tissue culturing. This may occur as a result of nutrient depletion, contamination etc..
After subculturing of callus and a period of incubation how should we know the callus is responding positively? Is browning of the callus a problem?
Dear Anju.....I think Yes, Browning of calli is a problem in its further regeneration. I suggest to keep on subculturing or optimize concentration of antioxidants being added in media. I donot know about your crop and objective, but you can definitely control phenolics otherwise process will be halted or slowed badly.
Yes, browning of your cultured tissue is a problem and certainly hinders with its growth performance. therefore it is advisable to get rid of it ASAP. Also, a growing callus is visually different and shows soft, friable cell masses or shoots/organs (whatever the case may be).
Dear Anju
Oxidative browning is a common and severe problem in plant tissue cultures including callus cultures of some plant species, where browning is caused by accumulation and oxidation of phenolic compounds. Brown callus results in poor growth, eventually leading to cell death. Intensity of browning in callus could be decreased by adding a combination of 20-100 mg/l ascorbic acid and 10-50 mg/l citric acid to the medium.
Seied Mehdi Miri I am using the N6 medium supplemented with 2mg/L of 2,4-D and 0.2 mg/L of Kinetin for rice callus culture, however the rice callus is getting brownish. how is it possible to add the ascorbic acid and citric acid to the medium?is it possible to add it to the medium while making? or should make stocks solutions of ascorbic acid and citric acid after autoclavin the N6 medium?
Dear Najib, before Seid replies I happened to read your message. I recommend addition of ascorbic acid and citric acid (u can make stock of 50-100 mg/ml, filter sterile it and use as per your requirement) in media after autoclave. Good luck
Dear Najib, as mentioned by Dr Bakhsh, you can prepare the ascorbic acid and citric acid solution and use it.
But, i strongly suugest you please replace the 2,4-D growth regulator for callusing in rice or any plant. Because, after dedifferentiation of explant by 2,4-D, it is very hard to induce totipoency of the explant (redifferentiation). and 2,4-D always induced compact callus. Therefore, regeneration may not be occur overthere.
It is my personel opinion that you may use either NAA or IAA of the same amount of like 2,4-D.
Good luck
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