Histological Profiling Over Time to Optimize Root Cell Type-Specific Reporter Lines for Cell Sorting.

Abstract

Cell type-specific marker lines expressing fluorophores such as GFP or GUS can be used as starting material from which single cell types can be isolated by fluorescence-activated cell sorting (FACS) and/or for the study of root development. Establishing the stability of these lines is an essential step prior to further study to ensure that marker expression and localization is stable over time and during environmental perturbations of interest to researchers applying these lines as treatments. Here, we detail the use of root cross sectioning to investigate marker expression throughout the length and width of the root using the model legume Medicago truncatula as an example. In order to deal with the fact that plant cell walls are highly autofluorescent, we also describe the usage of confocal microscopy to conduct a lambda scan to discriminate autofluorescence from marker molecule expression.