The construction and partial characterization of plasmids containing complementary DNA sequences to human calcitonin precursor polyprotein.
(1) Total poly(A)-containing RNA isolated from human thyroid medullary carcinoma tissue was shown to direct the synthesis in the wheat germ cell-free system of a major (Mr 21000) and several minor forms of human calcitonin precursor polyproteins. Evidence for processing of these precursor(s) by the wheat germ cell-free system is also presented. (2) A small complementary DNA (cDNA) plasmid library has been constructed in the PstI site of the plasmid pAT153, using total human thyroid medullary carcinoma poly(A)-containing RNA as the starting material. (3) Plasmids containing abundant cDNA sequences were selected by hybridization in situ, and two of these (ph T-B3 and phT-B6) were characterized by hybridization--translation and restriction analysis. Each was shown to contain human calcitonin precursor polyprotein cDNA sequences. (4) RNA blotting techniques demonstrate that the human calcitonin precursor polyprotein is encoded within a mRNA containing 1000 bases. (5) The results demonstrate that human calcitonin is synthesized as a precursor polyprotein.
D002116:Calcitonin / Q000096:biosynthesis* / Q000235:genetics; D002474:Cell-Free System; D004247:DNA / Q000235:genetics*; D005786:Gene Expression Regulation; D006801:Humans; D066298:In Vitro Techniques; D010957:Plasmids; D011061:Poly A / Q000378:metabolism; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000096:biosynthesis* / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D013964:Thyroid Neoplasms / Q000032:analysis
D011498:Protein Precursors; D012334:RNA, Neoplasm; D011061:Poly A; D002116:Calcitonin; D004247:DNA