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Abstract：
Although cell-free protein expression has been widely used for the synthesis of single proteins, cell-free synthetic biology has rapidly expanded to new, more complex applications. One such application is the prototyping or implementation of complex genetic networks involving the expression of multiple proteins at precise ratios, often from different plasmids. However, expression of multiple proteins from multiple plasmids may inadvertently result in unexpected, off-target changes to the levels of the proteins being expressed, a phenomenon termed plasmid crosstalk. Here, we show that the effects of plasmid crosstalk─even at the qualitative level of increases vs decreases in protein expression─depend on the concentration of plasmids in the reaction and the type of transcriptional machinery involved in the expression. This crosstalk can have a significant impact on genetic circuitry function and even interpretation of simple experimental results and thus should be taken into consideration during the development of cell-free applications.
摘要：
尽管无细胞蛋白质表达已广泛用于合成单一蛋白质，无细胞合成生物学迅速扩展到新的，更复杂的应用。一个这样的应用是复杂的遗传网络的原型设计或实现，涉及以精确的比例表达多种蛋白质。通常来自不同的质粒。然而,来自多个质粒的多种蛋白质的表达可能无意中导致意想不到的结果，表达的蛋白质水平的脱靶变化，一种称为质粒串扰的现象。这里,我们表明，即使在蛋白质表达增加与减少的定性水平上，质粒串扰的影响也取决于反应中质粒的浓度和参与表达的转录机制的类型。这种串扰可能会对遗传电路功能甚至简单实验结果的解释产生重大影响，因此在开发无细胞应用时应予以考虑。
