Plant growth and morphogenesis are determined by the mechanical properties of its cell walls. Using atomic force microscopy, we have characterized the dynamics of cell wall elasticity in different tissues in developing roots of several plant species. The elongation growth zone of roots of all species studied was distinguished by a reduced modulus of elasticity of most cell walls compared to the meristem or late elongation zone. Within the individual developmental zones of roots, there were also significant differences in the elasticity of the cell walls of the different tissues, thus identifying the tissues that limit root growth in the different species. In cereals, this is mainly the inner cortex, whereas in dicotyledons this function is performed by the outer tissues—rhizodermis and cortex. These differences result in a different behaviour of the roots of these species during longitudinal dissection. Modelling of longitudinal root dissection using measured properties confirmed the difference shown. Thus, the morphogenesis of monocotyledonous and dicotyledonous roots relies on different tissues as growth limiting, which should be taken into account when analyzing the localization of associated molecular events. At the same time, no matrix polysaccharide was found whose immunolabelling in type I or type II cell walls would predict their mechanical properties. However, assessment of the degree of anisotropy of cortical microtubules showed a striking correlation with the elasticity of the corresponding cell walls in all species studied.