Jaunin Françoise, Fakan Stanislav
Center of Electron Microscopy, University of Lausanne, 27 Bugnon, CH-1005 Lausanne, Switzerland.
J Cell Biochem. 2002;85(1):1-9.
The model of in situ DNA replication provided by immunofluorescence and confocal imaging is compared with observations obtained by electron microscopic studies. Discrepancies between both types of observations call into question the replication focus as a persistent nuclear structure and as a replication entity where DNA replication takes place. Most electron microscopic analyses reveal that replication sites are confined to dispersed chromatin areas at the periphery of condensed chromatin, and the distribution of replication factors exhibits the same localization pattern. Moreover, rapid migration of newly synthesized DNA from the replication sites towards the interior of condensed chromatin regions obviously takes place during S-phase. It implies modifications of replication domains, hardly detectable by fluorescence microscopy. The confrontation of different observations carried out at light microscopic or electron microscopic levels of resolution lead to a conclusion that a combination of in vivo fluorescence analysis with a subsequent ultrastructural investigation performed on the same cells will represent an optimal approach in future studies of nuclear functions in situ.
将免疫荧光和共聚焦成像提供的原位DNA复制模型与电子显微镜研究获得的观察结果进行比较。两种观察结果之间的差异对复制焦点作为一种持久的核结构以及作为DNA复制发生的复制实体提出了质疑。大多数电子显微镜分析表明，复制位点局限于浓缩染色质周边的分散染色质区域，并且复制因子的分布呈现相同的定位模式。此外，在S期明显发生了新合成的DNA从复制位点向浓缩染色质区域内部的快速迁移。这意味着复制结构域的修饰，而这在荧光显微镜下很难检测到。在光学显微镜或电子显微镜分辨率水平上进行的不同观察结果的对比得出结论，在未来原位核功能研究中，将体内荧光分析与随后对同一细胞进行的超微结构研究相结合将是一种最佳方法。
