Precise and reliable gene expression via standard transcription and translation initiation elements
Precise and reliable gene expression via standard transcription and translation initiation elements
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DOI：
10.1038/nmeth.2404
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发表时间：
2013-04-01
期刊：
影响因子：
48
通讯作者：
Endy, Drew
中科院分区：
文献类型：
--
作者：
Mutalik, Vivek K.;Guimaraes, Joao C.;Endy, Drew
无法可靠地预测遗传元件新组合的定量行为限制了生物系统的合理工程。我们开发了一种用于在大肠杆菌中控制转录和翻译起始的遗传元件的表达盒架构：转录元件编码共同的mRNA起始，翻译元件使用在许多自然系统中发现的重叠遗传基序。我们根据这些定义设计了组成型和阻遏物调节启动子以及翻译起始元件的文库。我们测量了每个文库的活性分布，并选择了共同导致在 1,000 倍观察到的动态范围内表达的元素。我们研究了策划元件的所有组合，证明任意基因都能在两倍相对目标表达窗口内可靠地表达，可靠性接近 93%。我们期望此处验证的遗传元件定义可以共同扩展，以创建公共领域标准生物部分的集合，以支持在基因组规模上对基因表达进行可靠的正向工程。
An inability to reliably predict quantitative behaviors for novel combinations of genetic elements limits the rational engineering of biological systems. We developed an expression cassette architecture for genetic elements controlling transcription and translation initiation in Escherichia coli: transcription elements encode a common mRNA start, and translation elements use an overlapping genetic motif found in many natural systems. We engineered libraries of constitutive and repressor-regulated promoters along with translation initiation elements following these definitions. We measured activity distributions for each library and selected elements that collectively resulted in expression across a 1,000-fold observed dynamic range. We studied all combinations of curated elements, demonstrating that arbitrary genes are reliably expressed to within twofold relative target expression windows with similar to 93% reliability. We expect the genetic element definitions validated here can be collectively expanded to create collections of public-domain standard biological parts that support reliable forward engineering of gene expression at genome scales.