47(P-9) Single cell analysis of plant metabolites using LASER micro sampling
Details
The analysis of bulk tissue samples gives the informations about solute and metabolite concentrations of all compartments and different cell types, and this do not reflect the fine scale patterns between adjacent cells or provide information about processes, that are regulated on the single cell level. To understand the physiology of a whole organism, it is necessary to understand the behavior of a single cell as its fundamental unit. Such investigations essentially require single cell or even subcellular analysis. Along with the progress of plant physiology, the reliable method for single cell sampling is sought-after to investigate the precise metabolic profiling of different cell types. Research on single cells of higher plants is relatively difficult, mainly because of their small size and rigid cell walls. Single cell sampling of plant cell is usually performed by a fine oil-filled glass microcapillary, mounted on a micromanipulator. However, this method is time consuming and rather needs high skill due to the hard and thick cell wall barrier. We developed a unique method of laser-assisted single cell sampling and applied this to metabolite analysis at the resolution of the individual cell. Some pico-litter cell content of GFP expressed transgenic torenia (Torenia hybrida.) were picked up from a single cell (epidermal and petal cell) using this method. The trans gene (GFP) mRNA was detected by RT-PCR, and the petal pigments i.e. peonidin-3,5-diglucoside and malvidin-3-glucoside-5-(p-coumaroyl)-glucoside were also identified by using nano flow LC-ESI MS/MS and MALDI TOF MS. This method would be one of the most powerful tools for studying metabolic profiling of individual plant cells.
