DNA in proximity to the site of replication is preferentially alkylated in S phase 10T1/2 cells treated with N-methyl-N-nitroso-urea.
DNA in proximity to the site of replication is preferentially alkylated in S phase 10T1/2 cells treated with N-methyl-N-nitroso-urea.
复制标题
在用 N-甲基-N-亚硝基-脲处理的 S 期 10T1/2 细胞中，复制位点附近的 DNA 优先被烷基化。
DOI：
10.1093/carcin/3.10.1119
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发表时间：
1982
期刊：
影响因子：
4.7
通讯作者：
Kaufman,DG
中科院分区：
文献类型：
--
作者：
Cordeiro-Stone,M;Topal,MD;Kaufman,DG
复制DNA比本体DNA更容易被N-甲基-N-亚硝基脲（MNU）（一种自发活性甲基化剂）修饰。这一结论得到了两种不同实验方法结果的支持。首先，在S期用MNU处理同步化的C3 H 10 T1/2克隆8细胞，并将处理期间复制的DNA与大量DNA分离。这是通过用限制酶消化纯化的DNA并将复制叉相关的DNA保留在硝酸纤维素滤膜中来完成的。其次，将同步化的C3 H 10 T1/2克隆8细胞暴露于5-溴脱氧尿苷和[3 H]MNU，并在CsCl梯度中分离密度标记的复制DNA。这两种方法都显示与复制DNA相关的每个核苷酸残基的[3 H]甲基加合物比随机甲基化预期的多2.6至5.0倍。这些实验是在低MNU浓度（0.018 - 0.115 mM）下进行的，该浓度不会引起10 T1/2细胞中DNA合成的任何可检测抑制或修复复制的刺激。
Replicating DNA is more susceptible to modification by N-methyl-N-nitrosourea (MNU), a spontaneously active methylating agent, than bulk DNA. This conclusion is supported by results from two different experimental approaches. First, synchronized C3H 10T1/2 clone 8 cells were treated in S phase with MNU and DNA replicated during the period of treatment was separated from bulk DNA. This was done by digesting the purified DNA with restriction enzymes and retaining the replication fork-associated DNA in nitrocellulose filters. Second, synchronized C3H 10T1/2 clone 8 cells were exposed to 5-bromodeoxyuridine and [3H]MNU and the density-labelled, replicated DNA was separated in CsCl gradients. Both methods show 2.6 to 5.0 times more [3H]methyl adducts per nucleotide residue associated with replicating DNA than that expected from random methylation. These experiments were done at low MNU concentrations (0.018 – 0.115 mM) that did not cause any detectable inhibition of DNA synthesis or stimulation of repair replication in 10T1/2 cells.