Multi-dimensional studies of synthetic genetic promoters enabled by microfluidic impact printing.
Multi-dimensional studies of synthetic genetic promoters enabled by microfluidic impact printing.
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通过微流体冲击印刷实现合成遗传启动子的多维研究
DOI：
10.1039/c7lc00382j
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发表时间：
2017-06-27
期刊：
影响因子：
6.1
通讯作者：
Pan T
中科院分区：
文献类型：
--
作者：
Fan J;Villarreal F;Weyers B;Ding Y;Tseng KH;Li J;Li B;Tan C;Pan T
天然遗传启动子受多种顺式和反式调控因子的调控。对于这些启动子的定量研究，通常只改变单一因子的浓度，以获得启动子相对于该因子的剂量响应或传递函数。这样的实验设计限制了我们理解启动子中多个调控因子之间定量、组合相互作用的能力。这种限制主要是由于多因子实验设计产生的难以处理的实验组合数量。为了克服这一主要限制，我们整合了冲击打印和无细胞系统，使遗传启动子的多维研究成为可能。我们首先提出了一种梯度打印系统，该系统由平行压电悬臂梁作为可扩展的致动器阵列组成，可产生100pL - 10nL范围内可调体积的液滴，这有助于在1μ l液滴的1 - 10,000倍范围内实现高精度的直接稀释。接下来，我们将应用该技术研究合成遗传启动子中三个调控因子之间的相互作用。最后，利用多参数、多维数据建立了基因调控模块的数学模型。我们的工作为使用无细胞系统和液滴打印进行合成遗传结构的多维研究开辟了新的前沿。建立了一个多参数梯度生成系统，用于合成遗传模块的多维、高通量、低消耗的定量分析。
Natural genetic promoters are regulated by multiple cis and trans regulatory factors. For quantitative studies of these promoters, the concentration of only a single factor is typically varied to obtain dose response or transfer function of the promoters with respect to the factor. Such design of experiments has limited our ability to understand quantitative, combinatorial interactions between multiple regulatory factors at promoters. The limitation is primarily due to the intractable number of experimental combinations that arise from multifactorial design of experiments. To overcome this major limitation, we integrate impact printing and cell-free systems to enable multi-dimensional studies of genetic promoters. We first present a gradient printing system which comprises parallel piezoelectric cantilever beams as a scalable actuator array to generate droplets with tunable volumes in the range of 100pL – 10nL, which facilitates highly accurate direct dilutions in the range of 1 – 10,000 fold in a 1μL drop. Next, we apply this technology to study interactions between three regulatory factors at a synthetic genetic promoter. Finally, a mathematical model of gene regulatory modules is established using the multi-parametric and multi-dimensional data. Our work creates a new frontier in the use of cell-free systems and droplet printing for multi-dimensional studies of synthetic genetic constructs. A multi-parametric gradient generation system has been established for multi-dimensional, high-throughput, and low-consumption quantitative analysis of a synthetic genetic module.
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影响因子：
6.1
作者：
Dutka, Filip;Opalski, Adam S.;Garstecki, Piotr
通讯作者：
Garstecki, Piotr
DOI：
10.1073/pnas.1106752109
发表时间：
2012-05-15
影响因子：
11.1
作者：
Leung, Kaston;Zahn, Hans;Hansen, Carl L.
通讯作者：
Hansen, Carl L.
影响因子：
3.4
作者：
Patterson, GH;Knobel, SM;Piston, DW
通讯作者：
Piston, DW
影响因子：
6.1
作者：
Fan J;Li B;Xing S;Pan T
通讯作者：
Pan T
影响因子：
8.4
作者：
Hodgman, C. Eric;Jewett, Michael C.
通讯作者：
Jewett, Michael C.