Enzyme-Free Ligation of 5'-Phosphorylated Oligodeoxynucleotides in a DNA Nanostructure
Enzyme-Free Ligation of 5'-Phosphorylated Oligodeoxynucleotides in a DNA Nanostructure
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DOI：
10.1002/cbdv.201700315
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发表时间：
2017-09-01
影响因子：
2.9
通讯作者：
Richert, Clemens
中科院分区：
文献类型：
--
作者：
Kramer, Markus;Richert, Clemens
多组分反应是困难的合成转化。对于DNA来说，有一个特殊的机会来排列折叠纳米结构中的多条链，以便它们被预先组织以给出特定的序列。DNA折纸结构中的多链反应先前已经使用光化学交联、1，3 '-偶极环加成或氨基磷酸酯形成反应进行。在这里，我们报告了在一个小的折纸片，产生的DNA链长度高达600个核苷酸在一个单一的步骤中，二甲酰亚胺驱动的磷酸二酯的形成。该方法使用具有5 '-末端磷酸的未修饰的寡脱氧核苷酸作为起始材料。与涉及线性双链体的酶促多链连接相比，通过凝胶电泳检测到，双酰亚胺驱动的连接产生较少的副产物。PCR扩增出全长600 mer的产物。
Multicomponent reactions are difficult synthetic transformations. For DNA, there is a special opportunity to align multiple strands in a folded nanostructure, so that they are preorganized to give a specific sequence. Multistrand reactions in DNA origami structures have previously been performed using photochemical crosslinking, 1,3'-diploar cycloadditions or phosphoramidate-forming reactions. Here we report carbodiimide-driven phosphodiester formation in a small origami sheet that produces DNA strands up to 600 nucleotides in length in a single step. The method uses otherwise unmodified oligodeoxynucleotides with a 5'-terminal phosphate as starting materials. Compared to an enzymatic multistrand ligation involving linear duplexes, the carbodiimide-driven ligation gave fewer side products, as detected by gel electrophoresis. The full-length 600mer product was successfully amplified by polymerase chain reaction.