I want to do Fluorescence Activated Cell Sorting on plant roots (based on GFP fluorescence) to collect material for RNA-seq.
I would like to select out specific cell type (defined by the expression of gene A) at a specific developmental stage (defined by the expression of gene B).
So I would like to find a way to activate GFP upon expression of both A and B (sort of the opposite of a bicistronic vector).
What is a good way to do this?
For FACS you will need strong expression of the GFP (or venusYFP) marker. I don't know whether you would be able to achieve this by such activation as you describe. An alternative could be to cut the region of the root that is at the desired developmental stage using a cell specific marker line as used for the Benfey dataset.
You want to use the expression of A and B to control the GFP (mark the cells you want). You should KI the GFP after the promoters of A and B. The question is either A or B expression will activate GFP.
plan A: You can use Cre-Loxp system (or other), use gene B control Cre and after Cre cut loxp then A activate GFP.
plan B: You can add other color, mark cells with two colors.
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