Endo-β-mannanase is a type of cell wall degrading enzyme and it plays varied roles in different developmental events and/or different plants. Evolutionary analysis of the gene family will provide an insight into gene function and speciation. In this study, genome-wide analysis and expression profiling of tomato and soybean endo-β-mannanase gene families were performed. We have identified 11 and 20 endo-β-mannanase genes in the tomato and soybean genomes, respectively. Sequence alignment revealed three α-helices (αA, αB, and αC) of plant endo-β-mannanases were conserved in tomato, soybean and Arabidopsis. Gene structure and phylogenetic analysis implied that loss of introns play an important role in the evolution of endo-β-mannanases. Evolutionary analysis of endo-β-mannanases from various plant species showed that they diversified before the division of lower and higher plants. Promoter sequence analysis and expression patterns analysis found that endo-β-mannanases are not only involving in growth and development process of tomato, but also associated with the stress response in plants.