I'm trying to obtain protoplasts from Physcomitrella patens. I have followed protocols from many literature papers. But I seem to get very few protoplasts. The moss that I'm using is very old (5 years) but I grind them very fine and plate them and after six- seven days try to isolate protoplasts from them. I have also prepared the Drieslase enzyme as per the protocols.
I have a theory that the mosses undergo transition from protonema to gametophyte in that seven days. I'm trying to see if there are any hormones or something that can maintain it in protonema stage.