Directed Evolution of Transcription Factor-Based Biosensors for Altered Effector Specificity.
Directed Evolution of Transcription Factor-Based Biosensors for Altered Effector Specificity.
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基于转录因子的生物传感器的定向进化以改变效应器特异性。
DOI：
10.1007/978-1-0716-2152-3_12
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发表时间：
2022
期刊：
影响因子：
--
通讯作者：
Machado LFM
中科院分区：
文献类型：
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作者：
Machado LFM
基于转录因子的生物传感器是合成生物学中用于感测工业上有价值的分子和临床上重要的代谢物的重要工具，因此在生物修复、工业生物技术和生物医学领域中呈现应用。以改变效应特异性为目的的变构转录因子（aTF）的定向进化具有开发新的生物传感器以检测天然和非天然分子的潜力，从而扩展了可用的基于aTF的生物传感器的范围。在本章中，我们描述了aTF定向进化的一般方法。库设计的理论进行了讨论，沿着与详细的方法，为改进的组合库的转换，并提出了实验搜索空间通过反选择使用荧光激活细胞分选（FACS）。
Transcription factor-based biosensors are important tools in Synthetic Biology for the sensing of industrially valuable molecules and clinically important metabolites, therefore presenting applications in the bioremediation, industrial biotechnology, and biomedical fields. The directed evolution of allosteric transcription factors (aTFs) with the aim of altering effector specificity has the potential for the development of new biosensors to detect natural and nonnatural molecules, expanding the scope of available aTF-based biosensors. In this chapter, we delineate a general method for the directed evolution of aTFs. The theory of library design is discussed, along with the detailed methodology for an improved transformation of combined libraries, and the experimental search space by counterselection using fluorescence-activated cell sorting (FACS) is presented.