Purification of pre-prolactin mRNA from bovine anterior pituitary glands

• Published in: The Journal of biological chemistry Vol. 254; no. 5; pp. 1516 - 1520
• Main Authors: Nilson, J H, Convey, E M, Rottman, F M
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 10.03.1979
• Subjects: Animals; Cattle; Molecular Weight; Pituitary Gland, Anterior - metabolism; Plants - metabolism; Poly A - analysis; Prolactin - biosynthesis; Protein Biosynthesis; Protein Precursors - biosynthesis; RNA, Messenger - isolation & purification; RNA, Messenger - metabolism; Triticum - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(17)37800-6

Total cytoplasmic RNA, isolated from bovine anterior pituitary glands, directed the synthesis of primarily two proteins in a wheat germ cell-free system. Analysis of wheat germ cell-free products by sodium dodecyl sulfate-slab gel electrophoresis, direct immunoprecipitation with prolactin aatibody, and prolactin- and growth hormone-specific radioimmunoassay indicated that the two proteins were pre-prolactin and pre-growth hormone. Agarose-urea-gel electrophoresis of poly(A)-containing RNA revealed the presence of a large symmetrical RNA peak with an estimated size of approximately 15 S. The RNA contained in this peak was eluted from the gel and translated in the wheat germ cell-free system. Analysis of the translation products by electrophoresis, direct immunoprecipitation, and radioimmunoassay resulted in the detection of only pre-prolactin. The extent of pre-prolactin mRNA purification was assessed by synthesizing DNA complementary to gel-purified pre-prolactin mRNA and then following the hybridization of this cDNA to gel purified pre-prolactin mRNA. The result of this hybridization is consistent with the cell-free translation studies, namely that pre-prolactin mRNA is the predominant mRNA component of pituitary poly(A)-containing RNA.