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首页> 外文期刊>Current Protocols in Nucleic Acid Chemistry >Automated Synthesis and Purification of Guanidine-Backbone Oligonucleotides
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Automated Synthesis and Purification of Guanidine-Backbone Oligonucleotides

机译：自动合成和净化Guanidine-Backbone寡核苷酸

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摘要

This protocol describes a method based on iodine and a base as mild coupling reagents to synthetize deoxyribonucleic guanidines (DNGs)— oligodeoxynucleotide analogues with a guanidine backbone. DNGs display unique properties, such as high cellular uptake with low toxicity and increased stability against nuclease degradation, but have been impeded in their development by the requirement for toxic and iterative manual synthesis protocols. The novel synthesis method reported here eliminates the need for the toxic mercuric chloride and pungent thiophenol that were critical to previous DNG synthesis methods and translates their synthesis to a MerMadeTM 12 automated oligonucleotide synthesizer. This method can be used to synthesize DNG strands up to 20 bases in length, along with 5/-DNG-DNA-3/ chimeras, at 1- to 5-mumol scales in a fully automated manner. We also present detailed and accessible instructions to adapt the MerMadeTM 12 oligonucleotide synthesizer to enable the parallel synthesis of DNG andDNA/RNA oligonucleotides. Because DNG linkages alter the overall charge of the oligonucleotides, we also describe purification strategies to generate oligonucleotides with varying lengths and numbers of DNGs, based on extraction or preparative-scale gelelectrophoresis, along with methods to characterize the final products. Overall, this article provides an overview of the synthesis, purification, and handling of DNGs and mixed-charge DNG-DNA oligonucleotides.
机译：这个协议描述了一个基于碘的方法和一个基地轻度耦合试剂合成脱氧核糖核胍(dng)用胍oligodeoxynucleotide类似物骨干。高与低毒性和细胞吸收增加了稳定性对核酸酶降解,但在其发展的阻碍有毒和迭代手册要求合成协议。这里不需要有毒的报道氯化汞和辛辣的苯硫酚以前DNG合成方法的关键吗和翻译MerMadeTM 12的合成自动化的寡核苷酸合成器。方法可用于合成DNG链长到20基地,随着5 / -DNG-DNA-3 /嵌合体,1 -在一个完全5-mumol尺度自动化的方式。访问指令适应MerMadeTM 12寡核苷酸合成器来启用的平行合成DNG andDNA / RNA寡核苷酸。寡核苷酸的总负责,我们也描述净化策略来生成寡核苷酸不同长度和数字dng,基于提取或制备规模gelelectrophoresis以及方法描述最终的产品。文章提供了一个综合的概述,dng和的净化和处理mixed-charge DNG-DNA寡核苷酸。

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