کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1358656 | 981352 | 2013 | 11 صفحه PDF | دانلود رایگان |
A fully automated chemical method for the parallel and high-throughput solid-phase synthesis of 5′-triphosphate and 5′-diphosphate oligonucleotides is described. The desired full-length oligonucleotides were first constructed using standard automated DNA/RNA solid-phase synthesis procedures. Then, on the same column and instrument, efficient implementation of an uninterrupted sequential cycle afforded the corresponding unmodified or chemically modified 5′-triphosphates and 5′-diphosphates. The method was readily translated into a scalable and high-throughput synthesis protocol compatible with the current DNA/RNA synthesizers yielding a large variety of unique 5′-polyphosphorylated oligonucleotides. Using this approach, we accomplished the synthesis of chemically modified 5′-triphosphate oligonucleotides that were annealed to form small-interfering RNAs (ppp-siRNAs), a potentially interesting class of novel RNAi therapeutic tools. The attachment of the 5′-triphosphate group to the passenger strand of a siRNA construct did not induce a significant improvement in the in vitro RNAi-mediated gene silencing activity nor a strong specific in vitro RIG-I activation. The reported method will enable the screening of many chemically modified ppp-siRNAs, resulting in a novel bi-functional RNAi therapeutic platform.
Automated parallel and high-throughput synthesis of 5′-triphosphate oligonucleotides enables the preparation of chemically modified 5′-triphosphate small interfering RNAs designed to address RNA interference gene knockdown and directed immunostimulation.Figure optionsDownload as PowerPoint slide
Journal: Bioorganic & Medicinal Chemistry - Volume 21, Issue 3, 1 February 2013, Pages 722–732