Mansson P E, Sugino A, Harris S E
Nucleic Acids Res. 1981 Feb 25;9(4):935-46. doi: 10.1093/nar/9.4.935.
The abundant class of poly(A+)RNA [poly(A+)RNA11S] from rat seminal vesicle was used to synthesize ds-cDNA11S. The ds-cDNA11S was inserted and cloned into the Pst I site of pBR-322 using E. coli RR1 as host. Colony filter hybridization and restriction mapping was used to demonstrate that a 620 NTP long insert in a plasmid clone (pSV2) represents the almost full length structural gene coding for a precursor to the seminal vesicle secretion protein IV (SVS IV). The entire insert was sequenced and the coding region was matched with the known amino acid sequence. Most of the signal peptide sequence was derived from the DNA sequence. The insert in pSV2 was labelled and used to study the effect of testosterone on the accumulation of mRNA SVS IV. Administration of testosterone to castrated rats resulted in the induction of mRNA SVS IV from a few molecules per cell to levels of over 100,000 after 96 h of hormone treatment.
来自大鼠精囊的大量多聚腺苷酸加尾RNA(poly(A+)RNA11S)被用于合成双链cDNA11S。双链cDNA11S被插入并克隆到以大肠杆菌RR1作为宿主的pBR - 322的Pst I位点。菌落滤膜杂交和限制性图谱分析用于证明质粒克隆(pSV2)中一个620个核苷酸长的插入片段代表了编码精囊分泌蛋白IV(SVS IV)前体的几乎全长的结构基因。对整个插入片段进行测序,并将编码区与已知的氨基酸序列进行比对。大部分信号肽序列来自DNA序列。用pSV2中的插入片段进行标记,以研究睾酮对mRNA SVS IV积累的影响。给去势大鼠注射睾酮,在激素处理96小时后,导致mRNA SVS IV从每个细胞中的几个分子诱导至超过100,000的水平。