A Toolkit for Rapid Modular Construction of Biological Circuits in Mammalian Cells.,ACS Synthetic Biology

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A Toolkit for Rapid Modular Construction of Biological Circuits in Mammalian Cells.
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2019-11-05 , DOI: 10.1021/acssynbio.9b00322
João Pedro Fonseca ₁ , Alain R Bonny ₁ , G Renuka Kumar ₂ , Andrew H Ng ₃ , Jason Town ₁ , Qiu Chang Wu ₄ , Elham Aslankoohi ₁ , Susan Y Chen ₁ , Galen Dods ₁ , Patrick Harrigan ₁ , Lindsey C Osimiri _{1,

5} , Amy L Kistler ₂ , Hana El-Samad _{1,

2}

Affiliation

The ability to rapidly assemble and prototype cellular circuits is vital for biological research and its applications in biotechnology and medicine. Current methods for the assembly of mammalian DNA circuits are laborious, slow, and expensive. Here we present the Mammalian ToolKit (MTK), a Golden Gate-based cloning toolkit for fast, reproducible, and versatile assembly of large DNA vectors and their implementation in mammalian models. The MTK consists of a curated library of characterized, modular parts that can be assembled into transcriptional units and further weaved into complex circuits. We showcase the capabilities of the MTK by using it to generate single-integration landing pads, create and deliver libraries of protein variants and sgRNAs, and iterate through dCas9-based prototype circuits. As a biological proof of concept, we demonstrate how the MTK can speed the generation of noninfectious viral circuits to enable rapid testing of pharmacological inhibitors of emerging viruses that pose a major threat to human health.

中文翻译：

在哺乳动物细胞中快速模块化构建生物回路的工具包。

快速组装和原型细胞电路的能力对于生物学研究及其在生物技术和医学中的应用至关重要。用于组装哺乳动物DNA电路的当前方法费力，缓慢且昂贵。在这里，我们介绍了Mammalian ToolKit（MTK），这是一种基于Golden Gate的克隆工具包，可用于大型DNA载体的快速，可复制和通用组装及其在哺乳动物模型中的实现。MTK由精选的，模块化的零件组成的精选库组成，这些零件可以组装成转录单位，然后进一步编织成复杂的电路。我们通过使用MTK生成单整合着陆垫，创建和交付蛋白质变体和sgRNA的库，以及通过基于dCas9的原型电路进行迭代来展示MTK的功能。作为概念的生物学证明，

更新日期：2019-11-05

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