In Situ Amplification‐Based Imaging of RNA in Living Cells

Qing, Zhihe; Xu, Jingyuan; Hu, Jinlei; Zheng, Jing; He, Lei; Zou, Zhen; Yang, Sheng; Tan, Weihong; Yang, Ronghua

doi:10.1002/ange.201812449

Cited by 46 publications

(14 citation statements)

References 131 publications

(309 reference statements)

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“…5 Some traditional approaches have been developed to detect miRNA, such as Northern blotting, 6 microassays, polymerase chain reaction (PCR), and fluorescence in situ hybridization. 7,8 Despite the significant success for the sensitive detection of miRNA, these methods can only work in vitro or fixed cells.…”

mentioning

confidence: 99%

Near-Infrared Light Controllable DNA Walker Driven by Endogenous Adenosine Triphosphate for in Situ Spatiotemporal Imaging of Intracellular MicroRNA

Kong

Zhang

et al. 2021

ACS Nano

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As a powerful signal amplification tool, the DNA walker has been widely applied to detect rare microRNA (miRNA) in vivo. Despite the significant advances, a nearinfrared (NIR) light controllable DNA walker for signal amplification powered by an endogenous initiator has not been realized, which is crucial for spatiotemporal imaging of miRNA in living cells with high sensitivity. Herein, we constructed a NIR-photoactivatable DNA walker system, which was powered by endogenous adenosine triphosphate (ATP) for in situ miRNA imaging with spatial and temporal resolution. The system was very stable with an extremely low fluorescent background for the bioimaging in living cells. We employed upconversion nanoparticles (UCNPs) as the carriers of the DNA probe and transducers of converting NIR to UV light. Coupled with the DNA walker fueled by intracellular ATP, a smart system based on the NIR light initiated DNA walker was successfully developed for precise spatiotemporal control in living cells. Triggered by NIR light, the DNA walker could autonomously and progressively travel along the track with the assistance of intracellular ATP. The system has been successfully applied for in situ miRNA imaging in different cell lines with highly spatial and temporal resolution. This strategy can expand NIR photocontrol the DNA walker for precise imaging in a biological system.

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confidence: 99%

Near-Infrared Light Controllable DNA Walker Driven by Endogenous Adenosine Triphosphate for in Situ Spatiotemporal Imaging of Intracellular MicroRNA

Kong

Zhang

et al. 2021

ACS Nano

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“…Based on target-specific CHA reactions, Tan’s group built a molecularly engineered, entropy-driven three-dimensional DNA amplifier for ultrasensitive detection of mRNAs and a cascade amplification nanoprobe for monitoring telomerase activity in living cells . Although enzyme-free CHA technology has improved the sensitivity of detection and found wide applications in bioanalysis and bioimaging, − researchers have continued to seek lower limits of detection to meet clinical requirements. Thus, a two-layer cascade employing upstream CHA circuits to initiate a second layer of hybridization chain reaction (HCR) circuits was developed, , allowing higher order amplification.…”

Section: Introductionmentioning

confidence: 99%

Highly Sensitive Detection of miR-21 through Target-Activated Catalytic Hairpin Assembly of X-Shaped DNA Nanostructures

Chai

et al. 2021

Anal. Chem.

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MicroRNAs (miRNAs) are found in extremely low concentrations in cells, so highly sensitive quantitation is a great challenge. Herein, a simple dual-amplification strategy involving target-activated catalytic hairpin assembly (CHA) coupled with multiple fluorophores concentrated on one X-shaped DNA is reported. In this strategy, four hairpin probes (H1, H2, H3, and H4) are modified with FAM and BHQ1 at both sticky ends, while a circulating hairpin probe (H0) is used to activate CHA circuits once it binds to complementary sequences in the target miR-21 (T). The powerful dual-amplification cascades in Förster resonance energy transfer (FRET)-based nonenzymatic nucleic acid circuits are triggered by T–H0-activated formation of the X-shaped DNA nanostructure, freeing T–H0 for the next CHA reaction cycle. CHA circuits increase the fluorescence due to the wide distance between FAM and BHQ1 in the formed X-shaped DNA nanostructure, resulting in signal amplification and highly sensitive detection of miR-21, with a limit of detection (LOD, 3σ) of 0.025 nM, which is 25.6 or 57.6 times lower than that obtained through a single-amplification strategy without multiple fluorophores on one X-shaped DNA or CHA circuit. Furthermore, this cascade reaction was completed in 45 min, effectively avoiding target degradation. This new enzyme-free signal amplification strategy holds promising potential for sensitively detecting different DNA or RNA sequences by simply adapting the fragment of the H0 sequence complementary to the target.

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“…At present, there are many methods for detecting SNPs (Uppu et al, 2016;Zhao et al, 2018;Prezza et al, 2019), including gene sequencing technology (Kim et al, 2016), capillary electrophoresis technology (Zhao et al, 2019), reversed-phase high performance liquid chromatography (HPLC) detection (Jones et al, 1999;Tonosaki et al, 2013), gene chip technology (Tu et al, 2018), etc. Although these methods can effectively detect SNPs, most of them PCR technology is required to amplify DNA (Qing et al, 2019), which is expensive and timeconsuming. Furthermore, the probability of obtaining a false positive is high and the process is complex (Matsuda, 2017).…”

Section: Introductionmentioning

confidence: 99%

Detection of Single Nucleotide Polymorphisms by Fluorescence Embedded Dye SYBR Green I Based on Graphene Oxide

Xia

Jing

et al. 2021

Front. Chem.

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The detection of single nucleotide polymorphisms (SNPs) is of great significance in the early diagnosis of diseases and the rational use of drugs. Thus, a novel biosensor based on the quenching effect of fluorescence-embedded SYBR Green I (SG) dye and graphene oxide (GO) was introduced in this study. The probe DNA forms a double helix structure with perfectly complementary DNA (pcDNA) and 15 single-base mismatch DNA (smDNA) respectively. SG is highly intercalated with perfectly complementary dsDNA (pc-dsDNA) and exhibits strong fluorescence emission. Single-base mismatch dsDNA (SNPs) has a loose double-stranded structure and exhibits poor SG intercalation and low fluorescence sensing. At this time, the sensor still showed poor SNP discrimination. GO has a strong effect on single-stranded DNA (ssDNA), which can reduce the fluorescence response of probe DNA and eliminate background interference. And competitively combined with ssDNA in SNPs, quenching the fluorescence of SG/SNP, while the fluorescence value of pc-dsDNA was retained, increasing the signal-to-noise ratio. At this time, the sensor has obtained excellent SNP resolution. Different SNPs detect different intensities of fluorescence in the near-infrared region to evaluate the sensor's identification of SNPs. The experimental parameters such as incubation time, incubation temperature and salt concentration were optimized. Under optimal conditions, 1 nM DNA with 0–10 nM linear range and differentiate 5% SNP were achieved. The detection method does not require labeling, is low cost, simple in operation, exhibits high SNP discrimination and can be distinguished by SNP at room temperature.

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In Situ Amplification‐Based Imaging of RNA in Living Cells

Cited by 46 publications

References 131 publications

Near-Infrared Light Controllable DNA Walker Driven by Endogenous Adenosine Triphosphate for in Situ Spatiotemporal Imaging of Intracellular MicroRNA

Near-Infrared Light Controllable DNA Walker Driven by Endogenous Adenosine Triphosphate for in Situ Spatiotemporal Imaging of Intracellular MicroRNA

Highly Sensitive Detection of miR-21 through Target-Activated Catalytic Hairpin Assembly of X-Shaped DNA Nanostructures

Detection of Single Nucleotide Polymorphisms by Fluorescence Embedded Dye SYBR Green I Based on Graphene Oxide

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