Cellobiose phosphorylase (EC 2.4.1.20, CBPase) is an enzyme that catalyzes the reversible phosphorolysis of cellobiose into α-D-glucose 1-phosphate (G-1-P) and D-glucose. As the acceptor specificity of the reverse reaction is broad, CBPase can synthesize hetero β-disaccharides from G-1-P and monosaccharides as acceptors. Here, in order to develop an efficient system for producing a rare β-disaccharide like 4- O -β-D-glucopyranosyl-D-xylopyranose (GX) using CBPase, two attempts were examined. Firstly, the CBPase gene from Cellvibrio gilvus was heterologously expressed as an intracellular enzyme using Aspergillus oryzae . The recombinant CBPase was enclosed in the mycelia of the host cell by cold acetone treatment and prepared as a mycelial dry powder. Secondly, to facilitate the synthesis of GX by removing glucose from the reaction mixture, a commercial dry yeast was added to the reaction mixture. The powdered CBPase-yeast combined system produced an approximately 3.5 times higher yield of GX from cellobiose than the reaction without yeast.