DNA AND RNA SEQUENCE DETERMINATION BASED ON PHOSPHOROTHIOATE CHEMISTRY
DNA AND RNA SEQUENCE DETERMINATION BASED ON PHOSPHOROTHIOATE CHEMISTRY
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DOI：
10.1126/science.2453926
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发表时间：
1988-06-10
期刊：
影响因子：
56.9
通讯作者：
ECKSTEIN, F
中科院分区：
文献类型：
--
作者：
GISH, G;ECKSTEIN, F
磷酸和硫代磷酸二酯之间的反应性差异是DNA和RNA测序的化学降解方案的基础。硫代磷酸酯基团在四个单独的酶促反应中掺入核酸中，其中三个是天然核苷三磷酸，一个是α-磷酸。硫代三磷酸在每个反应中。通过烷基化形成水解不稳定的硫代磷酸酯三酯来实现选择性链切割。作为一个例子，序列分析的M13噬菌体DNA和RNA的转录制备与SP 6 RNA聚合酶。
The difference in reactivity between phosphate and phosphorothioate diesters is the basis of a chemical degradation scheme for the sequencing of DNA and RNA. The phosphorothioate groups are incorporated into the nucleic acid in four separate enzymatic reactions, with three of the natural nucleoside triphosphates and one .alpha.-thiotriphosphate in each reaction. Selective strand cleavage is achieved through alkylation to form the hydrolytically labile phosphorothioate triester. As an example, the sequence analysis is presented of M13 phage DNA and of RNA prepared by transcription with SP6 RNA polymerase.