Department of Chemistry, Michigan Technological University, Houghton, Michigan.
College of Forest Resources and Environmental Science, Michigan Technological University, Houghton, Michigan.
Curr Protoc Nucleic Acid Chem. 2020 Sep;82(1):e111. doi: 10.1002/cpnc.111.
This protocol provides details for the preparation of nucleoside phosphoramidites with 1,3-dithian-2-yl-methyl (Dim) and 1,3-dithian-2-yl-methoxycarbonyl (Dmoc) as protecting groups, and a linker with Dmoc as the cleavable function, then using them for solid phase synthesis of sensitive oligodeoxynucleotides (ODNs). Using these Dim-Dmoc phosphoramidites and Dmoc linker, ODN synthesis can be achieved under typical conditions using phosphoramidite chemistry with slight modifications, and ODN deprotection and cleavage can be achieved under mild conditions involving oxidation with sodium periodate at pH 4 followed by aniline at pH 8. Under the mild deprotection and cleavage conditions, many sensitive functional groups including but not limited to esters, thioesters, alkyl halides, N-aryl amides, and α-chloroamides-which cannot survive the basic and nucleophilic deprotection and cleavage conditions such as concentrated ammonium hydroxide and dilute potassium methoxide used in typical ODN synthesis technologies-can survive. Thus, it is expected that the Dim-Dmoc ODN synthesis technology will find applications in the synthesis of ODNs that contain a wide range of sensitive functional groups. © 2020 Wiley Periodicals LLC. Basic Protocol: Synthesis, deprotection, cleavage, and purification of sensitive oligodeoxynucleotides Support Protocol 1: Synthesis of Dim-Dmoc nucleoside phosphoramidites Support Protocol 2: Preparation of CPG with a Dmoc linker Support Protocol 3: Synthesis of a phosphoramidite containing a sensitive alkyl ester group.
本方案提供了带有 1,3-二硫杂环戊烷-2-基甲基（Dim）和 1,3-二硫杂环戊烷-2-基甲氧羰基（Dmoc）作为保护基团的核苷亚磷酰胺的制备细节，以及带有 Dmoc 作为可裂解基团的连接子，然后将其用于敏感寡脱氧核苷酸（ODN）的固相合成。使用这些 Dim-Dmoc 亚磷酰胺和 Dmoc 连接子，可以在使用亚磷酰胺化学的典型条件下进行 ODN 合成，只需稍加修改，并且可以在温和条件下进行 ODN 的脱保护和裂解，涉及在 pH4 下用高碘酸钠氧化，然后在 pH8 下用苯胺。在温和的脱保护和裂解条件下，许多敏感的官能团包括但不限于酯、硫酯、烷基卤化物、N-芳基酰胺和α-氯酰胺——这些官能团不能耐受典型 ODN 合成技术中使用的浓氨和稀甲醇钾等碱性和亲核性脱保护和裂解条件——可以存活。因此，预计 Dim-Dmoc ODN 合成技术将在含有广泛敏感官能团的 ODN 合成中得到应用。©2020 威利父子公司。基本方案：敏感寡脱氧核苷酸的合成、脱保护、裂解和纯化支持方案 1：Dim-Dmoc 核苷亚磷酰胺的合成支持方案 2：带有 Dmoc 连接子的 CPG 的制备支持方案 3：含敏感烷基酯基团的磷酰胺的合成。
