Halacarid mites inhabit mainly in the bottom of the sea and their body lengths are approximately 0.5 mm. Few biological information about halacarids have been obtained. The phylogenetic study of halacarid mites has not been progressed and the phylogenetic relationship within the family Halacaridae is still unknown. Particularly there is no molecular study to infer phylogenetic relationship within the family. In the annual meeting of Acarological Society of Japan 2007, we developed the method for molecular analysis of 28S nuclear ribosomal gene in haracarid mites. Subsequently, we have further progressed the technique of the molecular analysis to infer the phylogenetic relationship of haracarids. The aim of the present study is to elucidate the phylogenetic position of the subfamily Rhombognathinae on the bases of molecular data. Halacarid specimens which were obtained from seaweeds and sandy segment at Japanese seacoasts, consist of eight genera in five subfamilies: Halacarinae (Halacarus sp., Agauopsis spp., Thalassarachna spp.), Halixodinae (Badyagaue sp., Agaue sp.), Copidgnathinae (Copidognathus spp.), Actacarinae (Actacarus spp.) and Rhombognathinae (Rhombognathus spp.). The template DNA was extracted and 28S nuclear ribosomal gene of the DNA was amplified by PCR reactions. And we determined approximately 400 base pair length of the gene. To construct phylogenetic trees by neighbor-joining method we calculated genetic distances of each operational taxonomic unit considering 1) transversions and 2) transitions and transversions based on Tamura - Nei model by using MEGA ver. 4 (Tamura et al. 2007). As a result, we got two trees both showing two main clusters. One of them includes only one subfamily Rhombognathinae and another consist of Halacarinae, Copidgnathnae, Actacarinae and Halixodinae. Rhombognathinae was greatly differentiated from the other subfamilies. Thus, Rhombognathinae possibly is a distinct taxon phylogenetically and this result supports the monophyletic nature obtained previously by morphological analysis.