Kempers-Veenstra A E, Musters W, Dekker A F, Klootwijk J, Planta R J
Biochemisch Laboratorium, Vrije Universiteit, Amsterdam, The Netherlands.
Curr Genet. 1985;10(4):253-60. doi: 10.1007/BF00365621.
Deletions in the promoter region of the 37S pre-rRNA operon in yeast were constructed and analysed in vivo using an artificial ribosomal minigene present on an extrachromosomal yeast vector. Sequences required for correct transcription initiation were found to be located between positions -192 and +15 relative to the start; a 5'-deletion down to position -133 reduces the transcription yield of the minigene at least five-fold. To allow detection of transcription of the minigene in isolated nuclei of yeast transformed with a minigene-bearing plasmid we attempted to increase the minigene copy number. The transcription yield in vivo appeared not to be proportional to the copy number but was found to be greatly enhanced when two or three minigenes are present in tandem. alpha-Amanitin sensitivity of transcription of these minigenes in isolated nuclei proved that RNA polymerase I is responsible for their transcription.
构建了酵母中37S前体rRNA操纵子启动子区域的缺失，并使用存在于酵母染色体外载体上的人工核糖体小基因在体内进行分析。发现正确转录起始所需的序列位于相对于起始位点的-192至+15位置之间；5'端缺失至-133位置会使小基因的转录产量至少降低五倍。为了检测用携带小基因的质粒转化的酵母分离细胞核中该小基因的转录情况，我们试图增加小基因的拷贝数。体内转录产量似乎与拷贝数不成正比，但当两个或三个小基因串联存在时，转录产量会大大提高。在分离细胞核中这些小基因转录对α-鹅膏蕈碱的敏感性证明RNA聚合酶I负责它们的转录。
