Synthetic Biology with an All E. coli TXTL System: Quantitative Characterization of Regulatory Elements and Gene Circuits
Synthetic Biology with an All E. coli TXTL System: Quantitative Characterization of Regulatory Elements and Gene Circuits
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DOI：
10.1007/978-1-4939-7795-6_4
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发表时间：
2018-01-01
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影响因子：
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通讯作者：
Noireaux, Vincent
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文献类型：
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作者：
Marshall, Ryan;Noireaux, Vincent
在过去的十年中，新一代无细胞转录-翻译（TXTL）系统已被设计用于新兴的多学科应用。依赖dna的体外蛋白质合成技术已被开发用于合成生物学、生物和化学工程以及生物物理学等定量学科的应用。除了在生物安全层面方便外，新的TXTL平台还具有用户友好性；更多的负担得起的;在转录水平上更加通用，TX曲目涵盖数百个部分；更强大，在批量和连续模式下蛋白质产量可达几毫克/毫升。因此，TXTL正在崛起为一种流行的研究工具，并被越来越多的研究团体使用。虽然越来越多的应用程序证明TXTL是可靠的，但获得适当的TXTL技能非常重要，特别是对于定量应用程序。TXTL已成为特别有用的快速原型遗传设备，从单一的调控元件到基本电路基序。在本章中，我们描述了开发适当的TXTL实践来表征这些遗传部分的基本程序。我们使用我们实验室开发的全大肠杆菌TXTL系统，现在由Arbor Biosciences以myTXTL的名称商业化。
Over the past decade, a new generation of cell-free transcription-translation (TXTL) systems has been devised for emerging multidisciplinary applications. The DNA-dependent in vitro protein synthesis technology has been developed to tackle applications in synthetic biology, biological and chemical engineering, as well as quantitative disciplines such as biophysics. In addition to being convenient at the biosafety level, the new TXTL platforms are user-friendly; more affordable; more versatile at the level of transcription, with a TX repertoire covering hundreds of parts; and more powerful, with protein production reaching a few mg/mL in batch and continuous modes. As a consequence, TXTL is rising up as a popular research tool and is used by a growing research community. While TXTL is proving reliable for an increasing number of applications, it is important to gain appropriate TXTL skills, especially for quantitative applications. TXTL has become particularly useful to rapidly prototype genetic devices, from single regulatory elements to elementary circuit motifs. In this chapter, we describe the basic procedures to develop appropriate TXTL practices for the characterization of such genetic parts. We use an all E. coli TXTL system developed in our lab, now commercialized by Arbor Biosciences under the name myTXTL.