[EN] UNNATURAL POLYMERASE SUBSTRATES THAT CAN SUSTAIN ENZYMATIC SYNTHESIS OF DOUBLE STRANDED NUCLEIC ACIDS FROM A NUCLEIC ACID TEMPLATE AND METHODS OF USE<br/>[FR] SUBSTRATS NON NATURELS POUR POLYMÉRASES POUVANT SUPPORTER LA SYNTHÈSE ENZYMATIQUE D'ACIDES NUCLÉIQUES DOUBLE BRIN À PARTIR D'UNE MATRICE D'ACIDE NUCLÉIQUE ET PROCÉDÉS D'UTILISATION
申请人:LIFE TECHNOLOGIES
公开号:WO2009134469A1
公开(公告)日:2009-11-05
Nucleotide analogs that can sustain the enzymatic synthesis of double-stranded nucleic acid from a nucleic template are described. The nucleotide analogs include: (i) a base selected from the group consisting of adenine, guanine, cytosine, thymine, uracil and their analogs; (ii) a label attached to the base or analog of the base via a cleavable linker; (iii) a deoxyribose; and (iv) one or more phosphate groups. The linker and/or the label inhibits template directed polymerase incorporation of a further nucleotide substrate onto an extended primer strand. In addition, cleavage of the linker leaves a residue attached to the base which is not present in the natural nucleotide and which does not inhibit extension of the primer strand. The nucleotide analogs can therefore be used as reversible terminators in sequencing by synthesis methods without blocking the 3' hydroxyl group. Methods of sequencing DNA using the substrates are also described.
描述了可以维持从核酸模板中酶催化合成双链核酸的核苷酸类似物。核苷酸类似物包括:(i)从腺嘌呤、鸟嘌呤、胞嘧啶、胸腺嘧啶、尿嘧啶及其类似物组成的碱基;(ii)通过可裂解连接物连接到碱基或碱基类似物上的标签;(iii)脱氧核糖;以及(iv)一个或多个磷酸基团。连接物和/或标签抑制模板定向聚合酶在扩展引物链上进一步催化合成核苷酸底物。此外,连接物的裂解留下一个残基连接到碱基上,该残基不在天然核苷酸中存在,也不会抑制引物链的扩展。因此,可以将核苷酸类似物用作合成中可逆的终止子,而不会阻塞3'羟基。还描述了使用这些底物测序DNA的方法。