THYMINE GLYCOL LESIONS TERMINATE CHAIN ELONGATION BY DNA-POLYMERASE-I INVITRO
THYMINE GLYCOL LESIONS TERMINATE CHAIN ELONGATION BY DNA-POLYMERASE-I INVITRO
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DOI：
10.1093/nar/14.2.737
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发表时间：
1986-01-24
影响因子：
14.9
通讯作者：
BEARDSLEY, GP
中科院分区：
文献类型：
--
作者：
CLARK, JM;BEARDSLEY, GP
用四氧化锇对噬菌体H13mp9的单链环状DNA进行化学修饰，以引入特异性顺式胸腺嘧啶乙二醇损伤，这是电离辐射产生的一种主要类型的DNA损伤。使用大肠杆菌I或T4 DNA聚合酶的大片段在受损和未受损的模板上延伸寡核苷酸引物。反应产物通过沿着DNA序列梯的电泳进行分析。在模板中与胸腺嘧啶碱基相对的位置处终止的受损模板上的合成。这些结果表明，顺式胸腺嘧啶乙二醇损伤的单链DNA构成块的DNA聚合酶在体外的合成。令人惊讶的是，复制停止后，正确的核苷酸，dAMP，插入对面的病变。这些结果表明，胸腺嘧啶乙二醇损伤的主要作用是抑制DNA合成，损伤不是一种有效的诱变剂。
Single-strand circular DNA from bacteriopbage H13mp9 was chemically modified with osmium tetroxide to introduce specificallycis-thymine glycol lesions, a major type of DNA damage produced by ionizing radiation. An oligonucleotide primer was extended on damaged and undamaged templates using either the large fragment ofE.colipol I or T4 DNA polymerase. The reaction products were analysed by electrophoresis alongside a DNA sequence ladder. Synthesis on the damaged templates terminated at positions opposite thymine bases in the template. These results indicate thatcis-thymine glycol lesions in single-strand DNA constitute blocks to synthesis by DNA polymerasesin vitro. Surprisingly, replication halts after the correct nucleotide, dAMP, is inserted opposite the lesion. These results imply that the primary effect of the thymine glycol lesion is suppression of DNA synthesis and that the lesion is not a potent mutagen.