Abstract
Prolactin mRNA has been isolated using immunochemical techniques. Initial experiments demonstrated that 125I-labeled prolactin antibody was able to bind to pituitary polysomes but not to liver polysomes, suggesting that the binding is specific. Prolactin-synthesizing polysomes were immunoprecipitated by reaction with antiprolactin followed by anti-antibody. Immunoprecipitated polysomal RNA was chromatographed on oligo(dT)-cellulose, and the poly(A) RNA was sedimented through a sucrose gradient. This procedure resulted in a 32-fold purification of prolactin mRNA as determined by translation in a mRNA-dependent reticulocyte lysate assay. Translation analysis also suggested that the isolated prolactin mRNA is greater than 95% pure. The molecular weight of prolactin mRNA determined by electrophoresis on agarose gels containing 10 mM methyl mercury hydroxide was 350,000. Purified prolactin mRNA was used to synthesize full-length cDNA by means of avian myeloblastosis reverse transcriptase. Use of this cDNA as a hydbridization probe demonstrated that estrogen is able to increase the concentration of prolactin mRNA sequences in the pituitary.
Original language | English (US) |
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Pages (from-to) | 854-859 |
Number of pages | 6 |
Journal | Journal of Biological Chemistry |
Volume | 255 |
Issue number | 3 |
State | Published - 1980 |
ASJC Scopus subject areas
- Biochemistry
- Molecular Biology
- Cell Biology