I need to get information about the number of chromosomes or DNA content.
I have seedlings from in vitro that were frozen at -20 degrees after liquidation of the culture. They are seedlings from controlled crosses between species.
Classical methods of counting metaphase chromosomes on slides did not work out.
We are considering flow cytometry, although we are concerned that frozen tissue will not be good enough for this.
Do you know any other methods, perhaps more molecular ?
We would appreciate any suggestions for useful protocols.
In this article four protocols for determining the nuclear DNA content of plants are described.
https://www.nature.com/articles/nprot.2007.310
Hi Aleksandra, I don't know whether you work with orchids (which is often your target group), but we have an experience with Flow Cytometry of deep-frozen (-70°C) tissues of Dactylorhiza. It worked unless the tissue thawed, it was thus necessary to chop it in the buffer immediatelly after removing from the freezer, and analyse as soon as possible. The results were suitable for interpretation of the ploidy level. The only challenge is the endoreplication, you must check how serious problem it is in seedlings.
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