<p>Bohemian knotweed (<i>Reynoutria</i> x <i>bohemica</i>) represents one of the worst invasive plant species in Europe and North America, spreading particularly along river banks by means of rhizomes as well as stem segments and having large impact on local biodiversity and state budgets. Since there is limited knowledge on plant regeneration efficiency from stem segments and its physiological basis in this hybrid, which are important for control strategies set up, we conducted a lab experiment and related stem segment/rhizome sugars and bud phytohormonal concentrations with the rooting success of floated segments, which were harvested in different stem position and phenological stage. The highest regeneration level after one month lasting cultivation was observed in segments from the middle and basal part of stems at the beginning of flowering stage (83.3 and 100%) with sprout/rootlet average dry weight of approx. 0.5/0.06 and 0.8/0.07 g, respectively. This happened after the indolyl-3-acetic acid bud level fell dramatically (to approx. 0.015 nmol g⁻¹ FW) and rhizome soluble/insoluble sugar concentrations were at the minimum (approx. 50 mg.g⁻¹ DW). Thus, depending on the phenological stage, plant regeneration from stems can reach relatively high efficiency, comparable to rhizomes. Released phyto-hormonal block of lateral bud growth and weak assimilate sinks are required. This knowledge needs to be taken into account in prevention of knotweeds spreading.</p>