Abstract：
A basic question of cell biology is how DNA folds to chromosome. A number of recently accumulated evidences have suggested that folding of chromosome proceeds tightly coupled with DNA replication progresses. Drug-induced PCC is a useful tool for visualization of the interphase nuclei, in particular, S-phase, as S-phase prematurely condensed chromosomes (S-phase PCC). Active replicating DNA is labeled directly with Cy3-dUTP by bead loading method, and then S-phase nuclei is immediately condensed prematurely by calyculin A to obtain S-phase PCC. Active replicating regions on S-PCC are observed under a scanning confocal microscope. Cy3-dUTP-labeled S-phase PCCs clearly reveal the drastic transitional change of chromosome formation through S-phase, starting from a \"cloudy nebula\" to numerous numbers of \"beads on a string\" and finally to \"striped arrays of banding structured chromosome\" known as G- or R-banding pattern. The number, distribution, and shape of replication foci were also measured in individual subphase of S-phase; maximally ~1400 foci of 0.35 μm average radius size were scored at the beginning of S-phase, and the number is reduced to ~100 at the end of S-phase. Drug-induced PCC clearly provided the new insight that eukaryote DNA replication is tightly coupled with the chromosome condensation/compaction for construction of eukaryote higher-ordered chromosome structure.
摘要：
细胞生物学的一个基本问题是DNA如何折叠成染色体。最近积累的许多证据表明，染色体的折叠与DNA复制的进展紧密相关。药物诱导的PCC是可视化相间核的有用工具，特别是,S相,作为S期过早缩合染色体（S期PCC）。活性复制DNA直接用Cy3-dUTP通过珠加载方法标记，然后，S相核立即被calyculinA过早凝聚，得到S相PCC。在扫描共聚焦显微镜下观察S-PCC上的活性复制区域。Cy3-dUTP标记的S期PCCs清楚地揭示了染色体形成通过S期的急剧过渡变化，从“浑浊的星云”开始到大量的“字符串上的珠子”，最后到“带状结构化染色体的条纹阵列”，称为G或R带模式。数字,分布，在S相的各个子相中还测量了复制灶的形状；在S相开始时对平均半径为0.35μm的最大〜1400个病灶进行评分，并且在S阶段结束时，该数量减少到〜100。药物诱导的PCC清楚地提供了新的见解，即真核生物DNA复制与染色体缩合/压缩紧密结合，以构建真核生物高阶染色体结构。
