Department of Chemistry, University of Oxford, Mansfield Road, Oxford OX1 3TA, U.K.
Department of Chemistry, University College London, 20 Gordon Street, London WC1H 0AJ, U.K.
J Am Chem Soc. 2023 May 3;145(17):9471-9480. doi: 10.1021/jacs.3c02350. Epub 2023 Apr 26.
Cell-free gene expression is a vital research tool to study biological systems in defined minimal environments and has promising applications in biotechnology. Developing methods to control DNA templates for cell-free expression will be important for precise regulation of complex biological pathways and use with synthetic cells, particularly using remote, nondamaging stimuli such as visible light. Here, we have synthesized blue light-activatable DNA parts that tightly regulate cell-free RNA and protein synthesis. We found that this blue light-activated DNA could initiate expression orthogonally to our previously generated ultraviolet (UV) light-activated DNA, which we used to generate a dual-wavelength light-controlled cell-free AND-gate. By encapsulating these orthogonal light-activated DNAs into synthetic cells, we used two overlapping patterns of blue and UV light to provide precise spatiotemporal control over the logic gate. Our blue and UV orthogonal light-activated DNAs will open the door for precise control of cell-free systems in biology and medicine.
无细胞基因表达是研究定义明确的最小环境中生物系统的重要研究工具，在生物技术中有广阔的应用前景。开发用于无细胞表达的 DNA 模板的控制方法对于复杂生物途径的精确调控和与合成细胞的使用非常重要，特别是使用远程、非损伤性的刺激，如可见光。在这里，我们合成了蓝光激活的 DNA 元件，可紧密调控无细胞 RNA 和蛋白质的合成。我们发现，这种蓝光激活的 DNA 可以与我们之前生成的紫外线（UV）光激活 DNA 正交启动表达，我们用它来生成双波长光控无细胞与门。通过将这些正交光激活 DNA 封装到合成细胞中，我们使用两种重叠的蓝光和 UV 光模式，对逻辑门进行精确的时空控制。我们的蓝光和 UV 正交光激活 DNA 将为生物学和医学中无细胞系统的精确控制打开大门。
