A nuclear scaffold fraction (designated P fraction elsewhere) comparable to a nuclear matrix was prepared from rat liver. This fraction was composed mainly of 45-49 kDa proteins and high-molecular-weight proteins (more than 90 kDa). In addition, a 370-bp repetitive sequence DNA fragment was derived predominantly from the EcoRI digest of the deproteinized P fraction. By an immunoblot affinity assay with the P fraction, the fragment was shown to have affinity for each of the 107- and 115-kDa proteins. Moreover, by a filter binding assay with a mixture of these proteins, the affinity level was estimated to be about 6 times as high in the native (double-stranded) fragment as in the denatured (single-stranded) fragment.