Hi,
I have some Catharantus roseus callus tissue (about 25 days on medium) that I kept in 12 hrs light condition on MS medium with 1 mg/L NAA and BAP. I don't want to let them differentiate and I need to maintain them in the undifferentiated mode. They are already green. Can I transfer them to dark now or is it too late?
Thank you
The differentiation seems to be already on its way. You can try low temperature treatment to slow the process. As suggested by Saeid.
As said, it is better to store the green callus in low temperature under dark to prevent further differentiation of cells.
Best wishes
you can place in the dark and at low temps but the chlorophyll mechanisms will take a very long time to degrade and any light source will bring things back to the future
As pointed out by others, greening suggests differentiation is already in process. My take is that you explore initiating another batch on same media but kept it in the dark throughout. You can later move the callus to light for greening and induction of morphogenesis. Reason is callus induction depends largely on the plant growth regulators rather then the presence/absence of light. All the best.
From my experience with other plant species, beside colour, important is the consistency of the calli.
Normally, a proliferative calli is also condensed, compacted and glassy, not friable.
I suggest - select these parts of calli and transfer them to a medium without growth regulators and maintain into darkness and lower temperature (5-10°C lower than previous conditions).
Good luck!
According my experience, I suggest to try pick up the white microcolonies from the green calli and transfer them to a medium without growth regulators (or with a 1/10 of obvious phytohormone conc.) and maintain into darkness and lower temperature as recommended the other. Good luck!
Select undifferentiated callus and transfer it on fresh medium under dark.
Catharanthus callus is highly recalcitrant to differentiation. Getting green never means the differentiation. You can batter maintain them on 1 mg/l 24-D and 0.1 mg/l BAP. This will only allow the healthy growth but no differentiation at al. Even no need to change light and temperature conditions.
I would try to reduce the amount of 2,4-D to a minimum (0,1 mg / l) and increase the amount of BAP to 1 mg / l. Maybe I would replace BAP with IBA.
Important is to induce embryogenesis in the dark, or organogenesis to light. Depends on how you want to get regeneration.
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