Koberna Karel, Ligasová Anna, Malínský Jan, Pliss Artem, Siegel Alan J, Cvacková Zuzana, Fidlerová Helena, Masata Martin, Fialová Markéta, Raska Ivan, Berezney Ronald
Department of Cell Biology, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Albertov 4, CZ-12800 Prague 2, Czech Republic.
J Cell Biochem. 2005 Jan 1;94(1):126-38. doi: 10.1002/jcb.20300.
DNA replication sites (RS) in synchronized HeLa cells have been studied at the electron microscopic level. Using an improved method for detection following the in vivo incorporation of biotin-16-deoxyuridine triphosphate, discrete RS, or foci are observed throughout the S-phase. In particular, the much larger RS or foci typically observed by fluorescence microscopic approaches in mid- and late-S-phase, are found to be composed of smaller discrete foci that are virtually identical in size to the RS observed in early-S-phase. Pulse-chase experiments demonstrate that the RS of early-S-phase are maintained when chased through S-phase and into the next cell generation. Stereologic analysis demonstrates that the relative number of smaller sized foci present at a given time remains constant from early through mid-S-phase with only a slight decrease in late-S-phase. 3-D reconstruction of serial sections reveals a network-like organization of the RS in early-S-phase and confirms that numerous smaller-sized replication foci comprise the larger RS characteristic of late-S-phase.
已在电子显微镜水平上研究了同步化的HeLa细胞中的DNA复制位点（RS）。使用一种改进的方法，在体内掺入生物素-16-脱氧尿苷三磷酸后进行检测，在整个S期观察到离散的RS或灶点。特别是，在S期中期和后期通过荧光显微镜方法通常观察到的大得多的RS或灶点，发现是由较小的离散灶点组成，其大小与在S期早期观察到的RS几乎相同。脉冲追踪实验表明，S期早期的RS在追踪到S期并进入下一代细胞时保持不变。体视学分析表明，从S期早期到中期，给定时间存在的较小尺寸灶点的相对数量保持恒定，在S期后期仅略有下降。连续切片的三维重建揭示了S期早期RS的网络状组织，并证实许多较小尺寸的复制灶点构成了S期后期特有的较大RS。
