mRNA for ovine follicle-stimulating hormone beta-chain. An in vitro translational assay

Polyadenylated mRNA from pituitary glands of castrate male sheep was used to demonstrate the presence of an mRNA coding for the beta-subunit of follicle-stimulating hormone (FSH). The mRNA fraction was translated in a wheat germ membrane-free system in the presence of [35S]cysteine. Affinity-purified antibodies, raised against the reduced and carbamylmethylated beta-subunit of ovine FSH (RCM-oFSH beta), were used to extract an in vitro translation product of Mr approximately equal to 15,000, as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Antibody binding of the product was readily blocked by unlabeled RCM-oFSH beta, but not by the RCM derivatives of the alpha- or beta-subunits of ovine luteinizing hormone. Tryptic peptide analysis using reversed phase high performance liquid chromatography yielded very similar patterns for the immunospecific translation product and authentic oFSH beta. FSH beta translation product was found to account for approximately 0.1-0.4% of the total isotope incorporation when mRNA preparations from castrate male sheep were used. Backgrounds for the immunoextraction procedure were kept at approximately 0.02% by using a unique two-cycle specific extraction method.