Plant Germline Development

Mário Luís da Costa, Ana Lúcia Lopes, Maria Isabel Amorim, Sílvia Coimbra

The arabinogalactan proteins (AGPs) are highly glycosylated proteins, ubiquitous in plants that have been linked to numerous aspects of sexual reproduction in several plant species, including the monoecious tree species Quercus suber. AGPs are found in cell membranes and cell walls of all types of tissues, including reproductive cells and organs. Pectins are cell wall components that also have been shown to change in composition and quantity during the maturations of the male and female gametophyte in cork oak. These findings were only possible to reveal, due to the histological study of AGP and pectins epitopes by immunolabeling. The immunofluorescence microscopy technique uses antibodies linked to fluorophores and relies on the specificity of the antibody binding to its antigen, labeling the epitope with a fluorescent dye.In the method presented here, we explore the immunolocalization technique performed in male and female flowers of Quercus suber, using London Resin (LR-White) as the embedding medium, after vacuum fixation with formaldehyde/glutaraldehyde. An extensive description of all the aspects of this technique is provided, from the plant material developmental stages selection to the critical analysis of results performed, continuously supported by troubleshooting recommendations.