Synthetic biology for fluorescent bioimaging

doi:10.12211/2096-8280.2021-060

Abstract

Abstract:

The rapid development of synthetic biology provides new opportunities for fluorescent bioimaging. Fluorescence imaging plays an important role in the visualization of biomolecules inside cells. Traditional visualization research methods have many disadvantages, such as the influence of excessive molecular weight of traditional fluorescent protein on the research object, and the low resolution of traditional fluorescence imaging observation system. The application of synthetic biology can develop new fluorescent nanomaterials, which have various advantages such as high stability, low toxicity and so on, and can be more widely used in cell internal visualization. Based on the principles of synthetic biology, we can establish new methods of material biosynthesis, develop fluorescent nanomaterials and probes with excellent performance, and develop new fluorescence imaging technologies. In the field of fluorescent biological imaging, synthetic biology mainly involves the design and synthesis of fluorescent materials, site-specific modification and labeling of biological target molecules, and controllable coupling of fluorescent probes and other macromolecules in different spatial relations. These novel fluorescent materials and molecular labeling techniques could be applied to molecular imaging and single particle tracking to explore intracellular molecular dynamic mechanisms. And new fluorescent nanomaterials developed in synthetic biology also can be used to tag different parts of viruses to trace the mechanism of their invasion so as to reveal pathogen infection and pathogenesis. This review summarizes advances of the synthetic biotechnology and the application in fluorescent imaging, including the synthesis of fluorescent nanomaterials and probes such as quantum dots, accurate labeling of proteins and nucleic acids, and the application in virus fluorescence imaging and tracing. We also discuss some existing problems and prospects in the field, such as controllable synthesis of fluorescent heterozygous biomaterials and multiple molecular labeling in situ. Synthetic biology has great development potential in the next decade. The multidisciplinary fusion of synthetic biology and fluorescence imaging technology will promote the development and progress of fluorescence imaging technology and expand the research field of biosynthesis.

Key words: synthetic biology, biological probes, molecular labeling, fluorescent imaging, virus tracing

摘要：

合成生物学的迅速发展为分子荧光标记与生物成像技术提供了新的机遇。基于合成生物学原理，可以建立材料生物合成新方法，开发性能优异的荧光纳米材料和探针，发展新的荧光成像技术。合成生物学应用于生物荧光成像，多涉及荧光材料与探针的设计合成、对生物靶标分子进行定点改造和修饰、荧光探针和靶标分子的可控时空耦合等以实现生物分子的精准特异性标记。这些荧光纳米材料和生物分子标记技术可应用于细胞内分子的荧光标记、成像和动态示踪，可视化解析相关的关键分子事件，从而深入揭示细胞内分子运动机制和病原致病机理等。本文主要综述了近年来合成生物学技术在生物荧光成像方面的应用，包括利用合成生物学技术合成量子点等荧光纳米材料与探针、对蛋白质和核酸分子的精准标记及其用于病毒荧光成像和示踪。最后，也对该领域面临的问题如荧光杂合生物材料可控合成、分子原位多重标记等进行了探讨和展望。合成生物学与荧光成像技术的交叉融合，将推动荧光成像技术发展和进步，并拓展合成生物学的研究领域。

关键词: 合成生物学, 生物探针, 分子标记, 荧光成像, 病毒示踪

CLC Number:

Q 81

Weihong WU, Wei LI, Xian’en ZHANG, Zongqiang CUI. Synthetic biology for fluorescent bioimaging[J]. Synthetic Biology Journal, 2022, 3(2): 369-384.

武伟红, 李炜, 张先恩, 崔宗强. 合成生物学与荧光成像技术[J]. 合成生物学, 2022, 3(2): 369-384.

Figures/Tables 12

Fig. 1 Controllable CdSe synthesis in yeast cells[45]

Fig. 2 CdTe biosynthesis in earthworm cells[50]

Fig. 3 CdSeZn and PrGd nanocrystals synthesized in E.coil cells [66]

Tab. 1 Properties of different fluorescent materials [74-76]

荧光材料	大小	荧光强度	荧光量子效率	荧光寿命
量子点	2～20 nm	+++++	0.1～0.85	20 min
荧光蛋白	4 nm	+	0.79	100 ms
荧光RNA	40 bp	+++	0.30～0.70	> 15 min

Fig. 4 Schematic diagram for imaging of Venus-TFFC(a) and mIrisFP-TFFC systems(b)[83-84]

Fig. 5 Schematic diagram for imaging polymerase activity in single cells through aptamer corn fluorophore[87](HEK293T cells transformed with the reporter plasmid could produce yellow fluorescence by adding fluorophore DFHO, but the fluorescence disappears after adding the RNA polymerase inhibitor actinomycin D.)

Fig. 6 Diagram for the working principle and structure diagram of bacterial riboswitch[89]

Fig. 7 Schematic diagram for imaging of the fluorescent RNA complex in HeLa cells[76]（A series of analogues of synthetic dye HBC combined with Pepper can emit fluorescence with different colors. HBC analogues were combined with Pepper in HeLa cells and are compared with mock cells. Imaging observation is carried out with confocal microscope and two-photon microscope.）

Fig. 8 Schematic diagram for HIV-1 entering cell through endocytosis[77]

Fig. 9 Real-time dynamic tracking of the influenza virus unpacking process[75][Under electron microscope, QD625 is black and round, while QD705 is triangular. vRNP is labeled with QD625 (red) and QD705 (green) respectively, and the results showed that QD625 and QD705 are released during the hulling process, respectively. IAV-released vRNPs released from IAV reach the nucleus through a three-stage active transport, where they undergo two diffusion modes.]

Fig. 10 TALEs labeled with quantum dots are used for gene composition imaging[95]

Fig. 11 SV40-QD VLPs for imaging atherosclerotic plaques in targeted mice[100](SV40 VLPs contain atherosclerotic targeting units, thrombin inhibitors, which can encapsulate as QD800. FH-QDS can target atherosclerotic areas in mice.)

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