Single-cell mRNA transfection studies: Delivery, kinetics and statistics by numbers
Single-cell mRNA transfection studies: Delivery, kinetics and statistics by numbers
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DOI：
10.1016/j.nano.2013.11.008
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发表时间：
2014-05-01
影响因子：
5.4
通讯作者：
Raedler, Joachim O.
中科院分区：
文献类型：
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作者：
Leonhardt, Carolin;Schwake, Gerlinde;Raedler, Joachim O.
在人工基因递送中，信使RNA（mRNA）是质粒DNA（pDNA）的有吸引力的替代物，因为它不需要转移到细胞核中。在这里，我们表明，与pDNA转染不同，mRNA介导的表达的传递统计和动力学是通用的和可预测的数学建模。我们使用延时显微镜和流式细胞术（FC）测量单细胞表达时间过程和增强型绿色荧光蛋白（eGFP）的水平。单细胞分析提供了对mRNA和eGFP的起始时间、寿命和表达率的分布的直接访问。我们介绍了一个两步随机传递模型，再现成功交付和翻译的mRNA分子的数量分布，从而剂量-反应关系。我们的研究结果建立了一个mRNA转染的统计框架，因此应该推进RNA载体和小干扰/微RNA为基础的药物的发展。来自临床编辑：这组作者建立了一个mRNA转染的统计框架，通过使用两步随机传递模型，再现成功传递和翻译的mRNA分子的数量分布，从而其剂量-反应关系。这项研究在理论和实验计划之间建立了良好的联系，并将有助于mRNA分子的细胞递送。(C)2014作者爱思唯尔公司出版All rights reserved.
In artificial gene delivery, messenger RNA (mRNA) is an attractive alternative to plasmid DNA (pDNA) since it does not require transfer into the cell nucleus. Here we show that, unlike for pDNA transfection, the delivery statistics and dynamics of mRNA-mediated expression are generic and predictable in terms of mathematical modeling. We measured the single-cell expression time-courses and levels of enhanced green fluorescent protein (eGFP) using time-lapse microscopy and flow cytometry (FC). The single-cell analysis provides direct access to the distribution of onset times, life times and expression rates of mRNA and eGFP. We introduce a two-step stochastic delivery model that reproduces the number distribution of successfully delivered and translated mRNA molecules and thereby the dose-response relation. Our results establish a statistical framework for mRNA transfection and as such should advance the development of RNA carriers and small interfering/micro RNA-based drugs.From the Clinical Editor: This team of authors established a statistical framework for mRNA transfection by using a two-step stochastic delivery model that reproduces the number distribution of successfully delivered and translated mRNA molecules and thereby their dose-response relation. This study establishes a nice connection between theory and experimental planning and will aid the cellular delivery of mRNA molecules. (C) 2014 The Authors. Published by Elsevier Inc. All rights reserved.