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The nitrogen-fixing root nodule is generally derived through a successful symbiotic interaction between legume plants and bacteria of the genus Rhizobium. A root nodule shelter hundreds of Rhizobia, which are thought to invade into the plant cells through an endocytosis-like process despite the existence of turgor pressure. Each invading Rhizobium is surrounded by the peribacteroid membrane to form the symbiosome, which results in the higher acquisition of host membrane materials. In this study, we show the localization of Rab11F, a RabA6b homolog with the large Rab-GTPase family, which was highly expressed in root nodules of Medicago sativa and M. truncatula. Rab11F-labeled organelles accumulated the membrane specific dye FM4-64 and were sensitive to Brefeldin A by forming aggregates after treatment with this drug. By co-localization with the cis-Golgi marker, GmMan1-mCherry, Rab11F-organelles formed tri-colored organelles, whereby Rab11F was located to the opposite side of GmMan1-mCherry indicating that Rab11F-labeled structures were localized within the trans-Golgi network (TGN). In root nodules, Rab11F was localized transiently at the infection thread-covering membrane on the side of infection droplets and the peribacteroid membranes. The symbiosome acquires Rab11F during the entry process and differentiation. However, the symbiosome did not recruit Rab11F after cessation of division. In conclusion, the legume plant seemed to use a specialized secretion pathway from the TGN, which was marked by Rab11F, to proliferate the symbiosome membrane.