McKnight S L, Hipskind R A, Reeder R
J Biol Chem. 1980 Aug 25;255(16):7907-11.
Xenopus laevis oocyte nuclei were manually isolated and incubated in vitro to synthesize RNA. Aliquots were removed before and during the reaction and prepared for electron microscopy, in order to observe the amplified ribosomal RNA (rRNA) genes. Before in vitro incubation the rRNA transcription units are covered with close-packed RNA polymerase molecules and attached rRNP fibers, which are interpreted to have initiated in vivo. With incubation, these close-packed transcription complexes move down the gene at a rate of 2.2 nucleotides per s. Behind this close-packed region we see RNA polymerase x nascent chain complexes at about one-fifth the in vivo frequency. These are interpreted to be in vitro initiation events. After 45 min of incubation, when most of the in vivo-initiated transcripts have terminated transcription, the in vitro-initiated transcripts define the same length transcription unit seen prior to incubation. These observations are consistent with correct in vitro initiation and termination by RNA polymerases on the endogenous amplified ribosomal DNA of Xenopus laevis.
非洲爪蟾卵母细胞核经手动分离后进行体外培养以合成RNA。在反应前及反应过程中取出等分试样并制备用于电子显微镜观察，以便观察核糖体RNA（rRNA）基因的扩增情况。在体外培养前，rRNA转录单位被紧密堆积的RNA聚合酶分子和附着的rRNP纤维覆盖，这些被认为是在体内起始的。随着培养的进行，这些紧密堆积的转录复合物以每秒2.2个核苷酸的速度沿着基因移动。在这个紧密堆积区域之后，我们看到RNA聚合酶与新生链复合物的出现频率约为体内频率的五分之一。这些被认为是体外起始事件。培养45分钟后，当大多数体内起始的转录本终止转录时，体外起始的转录本定义了与培养前所见相同长度的转录单位。这些观察结果与RNA聚合酶在非洲爪蟾内源性扩增核糖体DNA上进行正确的体外起始和终止相一致。
