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[与寡聚胸苷酸序列相邻的T7噬菌体DNA的互补性烷基化和切割]

[Complementarily addressed alkylation and cleavage of T7 phage DNA adjacent to the oligothymidylic sequences].

作者信息

Benimetskaia L Z, Gerasimova L M, Grineva N I, Karpova G G

出版信息

Mol Biol (Mosk). 1978 Sep-Oct;12(5):988-1001.

PMID:740002
Abstract

Effect of temperature and reagent excess on the alkylation of the T7 phage DNA by an alkylating derivative of hexaadenylate that containes a modifying group on the 3'-terminus has been studied. Under condition of saturation at 20 degrees the reagent covalently binds adjacent to the 133 5'-PuN2--4Tn greater than or equal to 4 sequences in the T7 DNA; at 40 degrees the reagent alkylates 24 5'-PuN2Tn greater than or equal to 6 sequences. DNA containing specific apurinic sites has been prepared due to elimination of the alkylated purines. This DNA has been cleaved at the apurinic sites and specific DNA fragments have been obtained. Three types of DNA fragments are formed due to alkylation at 20 degrees: one fragment with 26,500 nucleotides in length, seven fragments with 4500 nucleotides in length and 103--111 fragments with an average length of 190 nucleotides. Alkylation at 40 degrees with following apurinization and cleavage yields 21--23 fragments with the lengths: 1--31,500; 1--19,000; 3--9500 and 17--19 with 1100 nucleotides in average.

摘要

研究了温度和试剂过量对含有3'-末端修饰基团的六腺苷酸烷基化衍生物对T7噬菌体DNA进行烷基化反应的影响。在20℃饱和条件下,该试剂与T7 DNA中133个5'-PuN2--4Tn≥4序列相邻共价结合;在40℃时,该试剂使24个5'-PuN2Tn≥6序列烷基化。通过去除烷基化嘌呤制备了含有特定脱嘌呤位点的DNA。该DNA在脱嘌呤位点被切割,获得了特定的DNA片段。20℃烷基化产生三种类型的DNA片段:一种长度为26,500个核苷酸的片段,七个长度为4500个核苷酸的片段和103 - 111个平均长度为190个核苷酸的片段。40℃烷基化并随后进行脱嘌呤和切割产生21 - 23个片段,长度分别为:1 - 31,500;1 - 19,000;3 - 9500和17 - 19个平均长度为1100个核苷酸的片段。

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