THYMINE GLYCOLS AND UREA RESIDUES IN M13-DNA CONSTITUTE REPLICATIVE BLOCKS INVITRO
THYMINE GLYCOLS AND UREA RESIDUES IN M13-DNA CONSTITUTE REPLICATIVE BLOCKS INVITRO
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DOI：
10.1093/nar/13.22.8035
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发表时间：
1985-01-01
影响因子：
14.9
通讯作者：
WALLACE, SS
中科院分区：
文献类型：
--
作者：
IDE, H;KOW, YW;WALLACE, SS
用四氧化二锇（OsO4）处理M13 DNA，以浓度依赖的方式产生胸腺嘧啶二醇。为了形成含尿素的M13 DNA， oso4氧化的DNA在碱（pH12）中水解，将胸腺嘧啶二醇转化为尿素残基。对于含有胸腺嘧啶乙二醇和尿素的M13 DNA，大肠杆菌DNA聚合酶I know片段催化的DNA合成与模板DNA中存在的损伤数量成比例地减少。合成产物byE的测序凝胶分析。coliDNA聚合酶I和T4 DNA聚合酶显示DNA合成终止在假定的胸腺嘧啶乙二醇位点的反面，并在假定的尿素位点之前的一个核苷酸处终止。用锰代替反应混合物中的镁，增加了DNA合成的效率，从而使碱基与尿素相结合。对于含有胸腺嘧啶二醇的DNA，在锰存在下的处理能力强烈依赖于模板中胸腺嘧啶二醇的嘧啶5 '的存在。
Thymine glycols were produced in M13 DNA in a concentration dependent manner by treating the DNA with osmium tetroxide (OsO4). For the formation of urea-containing M13 DNA, OsO4-oxidized DNA was hydrolyzed in alkali (pH12) to convert the thymine glycols to urea residues. With both thymine glycol- and urea-containing M13 DNA, DNA synthesis catalyzed byEscherichia coliDNA polymerase I Klenow fragment was decreased in proportion to the number of damages present in the template DNA. Sequencing gel analysis of the products synthesized byE.coliDNA polymerase I and T4 DNA polymerase showed that DNA synthesis terminated opposite the putative thymine glycol site and at one nucleotide before the putative urea site. Substitution of manganese for magnesium in the reaction mix resulted in increased processivity of DNA synthesis so that a base was incorporated opposite urea. With thymine glycol-containing DNA, processivity in the presence of manganese was strongly dependent on the presence of a pyrimidine 5′ to the thymine glycol in the template.