The high resolution expression profiling method integrates spatial and temporal gene expression datasets across organ, development stage and tissue-specific cell types. Organs are dissected from whole plants and developmental zones can be microdissected along a developmental cline. Specific cell types are isolated by fluorescence-activated cell sorting (FACS) or laser capture microdissection (LCM). To isolate specific cell types by FACS, plant lines that express fluorescent markers in specific cell types are digested to release cells in suspension and then analyzed by flow cytometry to identify and collect the fluorescent cells-of-interest for transcript analysis. To isolate specific cell types by LCM, plant tissues are sectioned and analyzed by microscopy to identify cells-of-interest. Cells-of-interest are cut out of the sections by a precision laser and collected for transcript analysis.
Transcripts are analyzed by whole genome microarray or RNAseq platforms and integrated to generate an expression atlas of the plant. GrassRoots uses this rich dataset for gene expression element discovery: sequences from gene sets that are significantly expressed in all cell types and at all stages of development are used to develop truly constitutive gene expression elements and sequences from gene sets that have developmental stage or cell-type specific expression are used to design specific gene expression elements. In addition, GrassRoots conducts high resolution expression profiling on plants treated by environmental stress for trait gene discovery. The sensitivity of this dataset can detect novel stress response genes and the resolution can inform predictions of gene functions and testing strategies.