Triazole linking for preparation of a next-generation sequencing library from single-stranded DNA
Triazole linking for preparation of a next-generation sequencing library from single-stranded DNA
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DOI：
10.1093/nar/gky452
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发表时间：
2018-09-19
影响因子：
14.9
通讯作者：
Ito, Takashi
中科院分区：
文献类型：
--
作者：
Miura, Fumihito;Fujino, Tomoko;Ito, Takashi
下一代单链DNA测序技术正受到越来越多领域的关注。因此，正在积极测试各种方法来有效地对单链DNA进行接头标记。我们构思了一种新的化学酶方法，称为末端脱氧核苷酸转移酶(TDT)辅助，铜催化叠氮-炔环加成(CuAAC)介导的单链DNA连接(TCS连接)。在这种方法中，TDT被用来将单个3‘-叠氮化修饰的双脱氧核苷酸掺入到目标单链DNA的3’端，然后在CuAAC的介导下将叠氮修饰的3‘端连接到5’-乙炔化的合成接头上。这份报告介绍了TCS连接的第一个原理证明的应用，以及它在准备下一代测序文库中的使用。
Next-generation sequencing of single-stranded DNA (ssDNA) is attracting increased attention from a wide variety of research fields. Accordingly, various methods are actively being tested for the efficient adaptor-tagging of ssDNA. We conceived a novel chemoenzymatic method termed terminal deoxynucleotidyl transferase (TdT)-assisted, copper-catalyzed azide-alkyne cycloaddition (CuAAC)-mediated ssDNA ligation (TCS ligation). In this method, TdT is used to incorporate a single 3'-azide-modified dideoxyribonucleotide onto the 3'-end of target ssDNA, followed by CuAAC-mediated click ligation of the azide-incorporated 3'-end to a 5'-ethynylated synthetic adaptor. This report presents the first proof-of-principle application of TCS ligation with its use in the preparation of a next-generation sequencing library.