In vivo Analyse der Struktur von Chromosomenterritorien: Dyanamische Veränderungen während des Zellzyklus und der terminalen Differenzierung von Zellen

在活的哺乳动物细胞中，通过将胸腺嘧啶类似物与s期早期和中后期不同发射光谱的荧光团结合，可以对基因组DNA的富基因和贫基因部分进行差异标记。这两个部分代表有丝分裂染色体的R-带和G/ c带，并在间期核中形成“极化”染色体区域，基因贫乏且转录活性很大的G-灶聚集在核周围和核仁周围，而R-灶形成基因丰富且转录能力强的核内室（Sadoni等人，1999）。由于荧光标记染色单体和未标记染色单体的随机分离，在体内标记细胞核后出现了5-10个细胞周期，染色单体区域很少（Zink et al., 19998; 1999）。我们的方法首次允许在单个活细胞的水平上研究染色质排列及其动力学，双标记染色体。基于我们在第一个资助期内对高阶核结构的理解所取得的进展，我们将在第二个资助期内将研究重点放在细胞进入细胞周期或退出细胞周期进入GO或进行终端分化时，高阶核结构和有丝分裂染色体和染色体区域排列的变化问题上。

Gene rich and gene poor fractions of genomic DNA can be differentially labeled in living mammalian cells by the incorporation of thymidine analogs conjugated to fluorophores with different emission spectra during early and mid to late S-phase. These two fractions represent the R- and G/C-bands of mitotic chromosomes and form 'polarized' chromosome territories in interphase nuclei with gene poor and transcriptionally largely inactive G-foci clustered in the nuclear periphery and around the nucleoli, while R-foci form a gene rich and transcriptionally competent interior nuclear compartment (Sadoni et al., 1999). 5-10 cell cycles after in vivo labeling cell nuclei with few labeled chromatid territories emerge as a consequence of random segregation of fluorescently labeled and unlabeled chromatids (Zink et al., 19998; 1999). Our approach for the first time allows studies of chromatin arrangements and their dynamics in single living cells at the level of individual, double labeled chromosomes. Based on the progress we have achieved in our understanding of the higher order nuclear architecture during the first funding period, we will focus our studies during the second period on the question of changes of the higher order architecture and arrangements of mitotic chromosomes and chromosome territories, respectively, when cells proceed through the cell cycle or exit the cell cycle into GO or for terminal differentiation.