Separation of macromolecular RNAs by reversed-phase high-performance liquid chromatography

The reversed-phase high-performance liquid chromatography of synthetic and natural RNAs was studied. The results show that base composition alone cannot explain the retention characteristics of natural RNAs; secondary structures are probably involved. However, under the conditions described, the separation was independent of the sizes of the molecules. 18S and 28S ribosomal RNAs were eluted after tRNAs but before 9S globin mRNA. Furthermore, globin mRNA was resolved into two species, one containing and the other lacking poly(A). When applied to the separation of HeLa cell poly(A)-containing RNA, an heterogeneous pattern was obtained. Analysis of peptides synthesized by the mRNA showed that the separation is independent of size.