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We report a generalized methodology for the one-pot production of chemically modified functional RNA nanoparticles during in vitro transcription with T7 RNA polymerase. The efficiency of incorporation of 2'-fluoro-dNTP in the transcripts by the wild type T7 RNA polymerase dramatically increases in the presence of manganese ions, resulting in a high-yield production of chemically modified RNA nanoparticles functionalized with siRNAs that are resistant to nucleases from human blood serum. Moreover, the unpurified transcription mixture can be used for functional ex vivo pilot experiments.

Co-transcriptional Assembly of Chemically Modified RNA Nanoparticles Functionalized with siRNAs