Hernandez N, Keller W
Cell. 1983 Nov;35(1):89-99. doi: 10.1016/0092-8674(83)90211-8.
Using a whole cell extract of HeLa cells, we synthesized unspliced RNAs containing the first two leaders and the first intervening sequence of the adenovirus 2 major late transcription unit. Upon incubation of these pre-mRNAs in reaction mixtures containing a nuclear extract and a postnuclear fraction (S100), removal of the first intervening sequence and concomitant joining of the first leader to the second leader was observed. This splicing reaction requires proteins, Mg2+ ions, and ATP. The S100 fraction alone has no splicing activity but stimulates splicing when added to the nuclear extract. Upon fractionation of the postnuclear S100 by chromatography on ion exchange and gel filtration columns, the stimulatory activity copurifies with small ribonucleoprotein particles.
我们使用HeLa细胞的全细胞提取物，合成了包含腺病毒2型主要晚期转录单位的前两个前导序列和第一个内含序列的未剪接RNA。将这些前体mRNA在含有核提取物和核后组分（S100）的反应混合物中孵育后，观察到第一个内含序列的去除以及第一个前导序列与第二个前导序列的伴随连接。这种剪接反应需要蛋白质、镁离子和ATP。单独的S100组分没有剪接活性，但添加到核提取物中时会刺激剪接。通过离子交换色谱和凝胶过滤柱对核后S100进行分级分离后，刺激活性与小核糖核蛋白颗粒共纯化。
