I have been trying to establish new cell suspension cultures from some transgenic Arabidopsis (Ler) lines. After successfully obtaining calli from various tissues such as hypocotyls, shoot, root etc, we have been rather unsuccessful in converting these calli into a nice cell suspension cultures. To be a bit more specific:
a) Most suspension cultures I have worked with (but not established myself) were green in color. Ours are always brownish or dirty yellowish.
b) Growth rate is alright; however, the average number of cells in clumps is rather high. We have mostly visible cell clumps that are composed of more than 100 cells. They continue to 'pop-up' even after many rounds of selection for smaller cell clumps and single cell suspensions.
I have looked in as many guides as possible but could not find directions as how to solve these issues. Any comments/ideas/hints are more than welcomed. Thanks a lot,