The in vitro transcription (IVT) reaction used in the production of messenger RNA vaccines and therapies remains poorly quantitatively understood. Mechanistic modeling of IVT could inform reaction design, scale‐up, and control. In this work, we develop a mechanistic model of IVT to include nucleation and growth of magnesium pyrophosphate crystals and subsequent agglomeration of crystals and DNA. To help generalize this model to different constructs, a novel quantitative description is included for the rate of transcription as a function of target sequence length, DNA concentration, and T7 RNA polymerase concentration. The model explains previously unexplained trends in IVT data and quantitatively predicts the effect of adding the pyrophosphatase enzyme to the reaction system. The model is validated on additional literature data showing an ability to predict transcription rates as a function of RNA sequence length.

中文翻译：

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结合焦磷酸镁结晶效应的体外转录机制模型

用于生产信使 RNA 疫苗和疗法的体外转录 (IVT) 反应仍然缺乏定量的了解。 IVT 的机理建模可以为反应设计、放大和控制提供信息。在这项工作中，我们开发了 IVT 的机械模型，包括焦磷酸镁晶体的成核和生长以及随后晶体和 DNA 的团聚。为了帮助将该模型推广到不同的构建体，对转录速率作为靶序列长度、DNA 浓度和 T7 RNA 聚合酶浓度的函数进行了一种新颖的定量描述。该模型解释了 IVT 数据中先前无法解释的趋势，并定量预测了将焦磷酸酶添加到反应系统中的效果。该模型在其他文献数据上进行了验证，显示能够预测转录率作为 RNA 序列长度的函数。