Improvement of Self-amplifying RNA Quality and Potency with IVT Optimization
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Working with large/self-amplifying RNAs provides its own set of challenges. In this study we aimed to improve yields and quality of self-amplifying RNAs (saRNA) by optimizing conditions of the In Vitro Transcription (IVT) reaction. Most pre-made IVT buffers are not optimized for saRNA. We explored incubation temperatures, magnesium source and content, and reaction buffer conditions. These efforts have allowed us to produce RNAs as large as 16kb at high yields, low residual dsRNA, and high potencies.