Protein Immunolocalization in Maize Tissues

Abstract

The analysis of gene expression at transcript and at protein level is of outstanding importance in plant developmental biology. Proteins can be localized with subcellular resolution by immunolocalization using specific antibodies or generating reporter lines carrying the specific protein fused to a fluorescent protein. Immunolocalization is particularly suitable to confirm the expression pattern of transgenic reporter lines. It also represents a valid alternative, especially for plants such as maize, for which transformation is time consuming and still often unsuccessful, by-passing also some side-effects of fusion proteins. Indeed, the availability of specific antibodies for immunolocalizations and observations of maize tissues under a confocal microscope is a powerful tool for studying protein targeting to different cellular compartments.

In the following chapter we describe the complete procedure for the localization of proteins in different maize tissues both at cellular and sub-cellular level, using polyclonal or monoclonal antibodies. Tissues can be embedded in different substrates, such as paraplast, PEG400 and agarose, depending on the tissue and the desired use: epifluorescence or confocal microscope observations. An additional protocol for the analysis of the nuclear distribution of modified histones in squashed maize root apexes is also presented.