The genus Eucalyptus is composed of more than 600 species with high phenotypic and adaptation variability, allowing for different utilization of its wood. Three species are among the most planted: E. globulus, E. grandis and E. urophylla, They contain remarkable differences in lignin content, syringyl/guayacyl (S/G) ratio, cellulose productivity and adaptability to tropical conditions. Despite the importance of these species, information about gene expression and cell wall composition are still incomplete, impairing our ability to compare and comprehend the differences between these three species. The goal of the present work was to investigate the glycan diversity and content of these three Eucalyptus species using a new high-throughput approach named Glycome Proﬁling, which uses a large and diverse collection of 156 plant glycan-directed monoclonal antibodies. The results were combined with molecular data produced by xylem RNAseq transcriptome analyses. Our results suggest that signiﬁcant differences in cell wall composition and structure occur among the three species. The high glucose yield obtained from E. globulus is probably a result of different connections among cell wall molecules that allows facile degradation of cellulose. The observed differences can be, partly, explained by differential gene expression of cell wall related genes. These are pioneering data to provide an overview of glycan content and distribution in Eucalyptus and also a transcriptome comparison for these three species. Understanding the process of wood formation, composition and structure of cell wall components is critical for planning genetic improvement programs to develop less recalcitrant and higher yielding bioenergy feedstocks.