Stefanovsky V, Dimitrov S, Russanova V, Pashev I
Institute of Molecular Biology, Bulgarian Academy of Sciences, Sofia.
Mol Biol Rep. 1990 Nov;14(4):231-5. doi: 10.1007/BF00429890.
The presence of histones H1 and H4 at the sites of actual DNA synthesis has been studied with Ehrlich ascites tumour cells, pulse labeled for different times with 3H-thymidine and then treated with formaldehyde to crosslink histones to DNA. The fixed chromatin fragments were sonicated to reduce the size of DNA, purified in a CsCl gradient and immunoprecipitated with antibodies to histones H1 and H4. Determination of specific radioactivity in precipitated probes showed that both histones have been associated with nascent DNA even upon 1 min pulse with 3H-thymidine, thus indicating their presence near the replication fork.
利用艾氏腹水瘤细胞研究了实际DNA合成位点处组蛋白H1和H4的存在情况。用³H-胸腺嘧啶核苷对细胞进行不同时间的脉冲标记,然后用甲醛处理以使组蛋白与DNA交联。将固定的染色质片段进行超声处理以减小DNA的大小,在氯化铯梯度中纯化,并使用抗组蛋白H1和H4的抗体进行免疫沉淀。对沉淀探针中比放射性的测定表明,即使在用³H-胸腺嘧啶核苷脉冲标记1分钟后,这两种组蛋白也已与新生DNA相关联,从而表明它们存在于复制叉附近。