RNase H2 ofSaccharomyces cerevisiaeconsists of three essential subunits (Rnh201, Rnh202 and Rnh203) and plays a critical role in the removal of RNA incorporated in duplex DNA. In the present study, we purified indi-vidual subunits and heterodimeric subcomplexes to examine the assembly and biochemical function of subunits of RNase H2in vitro.