Wingender E, Frank R, Blöcker H, Wang L R, Jahn D, Seifart K H
Gesellschaft für Biotechnologische Forschung, Braunschweig, F.R.G.
Gene. 1988 Apr 15;64(1):77-85. doi: 10.1016/0378-1119(88)90482-9.
The gene coding for the major human ribosomal 5S RNA was chemically synthesized and cloned into a pUC13 vector. This approach was taken, because attempts to isolate the human 5S gene have thus far yielded either pseudogenes or variant 5S genes of unknown function. The synthetic human gene was transcribed by RNA polymerase III either in a crude HeLa cell extract or in a system reconstituted from partially purified transcription factors. Comparative studies with the Xenopus laevis somatic 5S gene show that the human gene is transcribed with similar fidelity and an efficiency of about 80% under optimal conditions. The time-course of transcription and optimal concentrations of template and transcription factors were found to be similar for both genes studied. The synthetic gene described may prove useful to study its interaction with human transcription factors in a homologous system.
编码主要人类核糖体5S RNA的基因经化学合成后克隆到pUC13载体中。之所以采用这种方法，是因为迄今为止，分离人类5S基因的尝试要么得到假基因，要么得到功能未知的变异5S基因。合成的人类基因由RNA聚合酶III在粗制的HeLa细胞提取物中或在由部分纯化的转录因子重构的系统中进行转录。与非洲爪蟾体细胞5S基因的比较研究表明，在最佳条件下，人类基因以相似的保真度转录，效率约为80%。研究发现，两个基因的转录时间进程以及模板和转录因子的最佳浓度相似。所述的合成基因可能被证明有助于在同源系统中研究其与人类转录因子的相互作用。
