Djeliova Vera, Russev George, Anachkova Boyka
Institute of Molecular Biology, Bulgarian Academy of Sciences, Sofia, Bulgaria.
J Cell Biochem. 2002;87(3):279-83. doi: 10.1002/jcb.10298.
HeLa cells were synchronized at late G1, early S, and late S phase of the cell cycle by nocodazole treatment. The cells were permeabilized with Triton X-100, digested with DNAse I, and extracted with 0.2 M ammonium sulfate to remove the digested chromatin. DNA was isolated from the residual chromatin attached to the nuclear matrix, digested with Hind III, and subjected to hybridization with [(32)P] labeled probe located upstream of the core region of the human beta-globin replication origin. The hybridization pattern revealed the existence of a DNase I sensitive site in the core region of the beta-globin replicator. The results suggest that association with the nuclear matrix induce alteration in the chromatin structure of the origin of replication that represents a more open chromatin configuration.
通过诺考达唑处理,将HeLa细胞同步于细胞周期的G1晚期、S早期和S晚期。用Triton X - 100使细胞通透,用DNA酶I消化,并用0.2M硫酸铵提取以去除消化的染色质。从附着于核基质的残留染色质中分离DNA,用Hind III消化,并用位于人β - 珠蛋白复制起点核心区域上游的[(32)P]标记探针进行杂交。杂交模式揭示了β - 珠蛋白复制子核心区域存在一个DNA酶I敏感位点。结果表明,与核基质的结合诱导了复制起点染色质结构的改变,这代表了一种更开放的染色质构型。