Kwiatkowski M, Nilsson M, Landegren U
Department of Medical Genetics, Uppsala University, Sweden.
Nucleic Acids Res. 1996 Dec 1;24(23):4632-8. doi: 10.1093/nar/24.23.4632.
The synthesis of oligodeoxynucleotides is marred by several problems that contribute to the formation of defective molecules. This in turn seriously limits the usefulness of such reagents in DNA diagnostics, molecular cloning, DNA structural analysis and in antisense therapy. In particular, depurination reactions during the cyclical steps of synthesis lead to strand scission during cleavage of the completed molecules from the support. Here we present a remedy to this problem: a novel disiloxyl linkage that connects oligonucleotides to the support withstands reaction conditions that allow the removal of the 5' parts of any depurinated molecules. This ensures that all molecules that preserve the 5' protecting group when cleaved from the support will have both correct 3'- and 5'-ends. We demonstrate the application of the support for synthesis of padlock probe molecules.
寡脱氧核苷酸的合成存在若干问题，这些问题会导致有缺陷分子的形成。这反过来又严重限制了此类试剂在DNA诊断、分子克隆、DNA结构分析及反义治疗中的用途。特别是，合成循环步骤中的脱嘌呤反应会导致在将完整分子从载体上切割下来时发生链断裂。在此，我们提出了针对这一问题的解决方法：一种将寡核苷酸连接到载体上的新型二硅氧烷基键，它能够承受允许去除任何脱嘌呤分子5'部分的反应条件。这确保了从载体上切割下来时保留5'保护基团的所有分子都将具有正确的3'端和5'端。我们展示了该载体在合成锁式探针分子中的应用。
