STRUCTURE OF INTERPHASE AND METAPHASE CHROMOSOMES

我们将继续分析催化的染色质组装反应 采用非洲爪哇卵母细胞(S-150)高速培养上清液。命运 反应中的组蛋白和非组蛋白的比例将是 特异组蛋白抗体免疫印迹和T25I的监测 新生染色质中DNA结合蛋白的标记。我们会 对新生的染色质的组成进行平行研究 非洲爪蛙肾细胞系的体外实验研究 我们所描述的组装过程反映了体内的 在染色质复制过程中发生的反应。 我们将刻画动态(扭转张力)，转录 活性染色质与阳性转录因子TFIIIA组装 5S核糖核酸基因。动态微染色体的组成将是 用组蛋白抗体免疫印迹和125I标记进行检测。 动态的、“开放的”核小体将以电子为特征 镜检和MNase(微球菌核酸酶)和DNase I酶消化。 我们将继续提纯组装所需的因素 转录活性的染色质。特别是，我们应该净化和 描述真核DNA旋转酶，一种在 我国扭转应变超级钢卷的生产与维护 动态染色质。这些研究应该能阐明 控制真核生物的基因表达。

We shall continue our analysis of the chromatin assembly reaction catalyzed by the high speed supernatant of Xenopus laevis oocytes (S-150). The fate of the histones and non-histone proteins during the reaction will be monitored by immunoblots with specific histone antibodies and by T25I labelling of the DNA-bound proteins in the nascent chromatin. We shall conduct parallel studies on the composition of nascent chromatin in a kidney cell line of Xenopus laevis, to ascertain whether the in vitro process of assembly which we have characterized reflects the in vivo reaction that takes place during chromatin replication. We will characterize the dynamic (torsionally strained), transcriptionally active chromatin assembled with TFIIIA, the positive transcription factor of the 5S RNA gene. The composition of the dynamic minichromosomes will be examined by immunoblots with the histone antibodies and by 125I labeling. The dynamic, "open" nucleosomes will be characterized by electron microscopy and by MNase (micrococcal nuclease) and DNase I digestion. We will continue the purification of the factors required for the assembly of transcriptionally active chromatin. In particular, we shall purify and characterize the eucaryotic DNA gyrase, an enzyme which plays a key role in the production and maintenance of the torsionally strained supercoils in dynamic chromatin. These studies should illuminate the mechanisms which govern gene expression in eucaryotes.