Enrichment of special Novikoff hepatoma and regenerating liver mRNA by hybridization to cDNA-cellulose

Abstract

Total polysomal poly(A)+ RNA of Novikoff hepatoma and of 18-hr regenerating rat liver were compared by analysis of their in vitro translational products on two-dimensional isofocusing/sodium dodecyl sulfate gels. This technique resolved the translated proteins sufficiently to permit detection of quantitative and some qualitative differences between the two mRNA populations. Excess cDNA from regenerating liver or Novikoff hepatoma, covalently linked to cellulose, was used to adsorb the complementary mRNA sequences from Novikoff hepatoma or regenerating liver. As shown by two-dimensional gel electrophoresis, the translated products of the bound mRNA fractions contained proteins common to both tissues. Novikoff hepatoma mRNA which did not bind to regenerating liver cDNA was enriched in sequences encoding for proteins 11/5.1, 15/6.8, 40/8.2, and 65/5.1 (shown as molecular weight/pI). These polypeptides were not detectable in the translational products of regenerating liver mRNA. Regenerating liver mRNA that was not bound to Novikoff hepatoma cDNA was enriched in sequences coding for proteins 12.5/4.9, 13.5/7.4, 17/8.2, 24/5.5, and 46/6.4; these proteins were not found in the translational pattern from Novikoff hepatoma. These results show that adsorption of mRNA to solid-phase cDNA provides a valuable technique for differentiating mRNA species in related tissues and for corresponding enrichment of these specific mRNAs.