Specific and sensitive detection of CircRNA based on netlike hybridization chain reaction

Dong, Jiani; Zeng, Zhuoer; Sun, Ruowei; Zhang, Xun; Zhang, Cheng; Chen, Chuanpin; Zhu, Qubo

doi:10.1016/j.bios.2021.113508

Cited by 36 publications

(25 citation statements)

References 44 publications

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“…(2) In comparison with the reported isothermal amplifications, , the TdT-mediated G-quadruplex synthesis endows this biosensor with excellent simplicity without the involvement of any DNA templates and exogenous primers. (3) The use of G-quadruplex/ThT complexes as a signal producer does not require specific labeled probes, enabling label-free detection of circRNA. (4) This assay may be implemented isothermally without the involvement of expensive complicated instruments .…”

Section: Discussionmentioning

confidence: 99%

“…Available technologies for the detection of circRNAs include microarray analysis, RNA sequencing, Northern blotting, , reverse transcription polymerase chain reaction (PCR), − droplet digital PCR, ligation-based PCR, and electrochemical measurement. , However, they usually suffer from some limitations such as the requirement of specialized equipment (for RNA sequencing and droplet digital PCR), ample computational power (for RNA sequencing), radioactively labeled probes (for Northern blotting), divergent primers design, and high-precision temperature control (for PCR), labor-intensive (for microarray analysis, RNA sequencing, and Northern blotting), and low sensitivity (for Northern blotting, ligation-based PCR, and electrochemical measurement). − Alternatively, isothermal amplification approaches including hybridization chain reaction, rolling circle amplification, , and cyclic enzymatic amplification have shown great potential in circRNA assay. Despite their advantages of isothermal and high amplification efficiency without the need for tedious procedures, − they often require specific reverse transcription primers, DNA templates, and fluorophore/quencher-labeled hairpin probes. − Consequently, new highly efficient methods for simple and sensitive analysis of circRNA biomarker in real samples are highly desirable.…”

mentioning

confidence: 99%

“…25,26 However, they usually suffer from some limitations such as the requirement of specialized equipment (for RNA sequencing and droplet digital PCR), ample computational power (for RNA sequencing), radioactively labeled probes (for Northern blotting), divergent primers design, and high-precision temperature control (for PCR), labor-intensive (for microarray analysis, RNA sequencing, and Northern blotting), and low sensitivity (for Northern blotting, ligation-based PCR, and electrochemical measurement). 19−26 Alternatively, isothermal amplification approaches including hybridization chain reaction, 27 rolling circle amplification, 28,29 and cyclic enzymatic amplification 30 have shown great potential in circRNA assay. Despite their advantages of isothermal and high amplification efficiency without the need for tedious procedures, 31−34 and fluorophore/quencher-labeled hairpin probes.…”

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confidence: 99%

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Target-Initiated Cascade Signal Amplification Lights up a G-Quadruplex for a Label-Free Detection of Circular Ribonucleic Acids

Wei

et al. 2022

Anal. Chem.

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Circular ribonucleic acids (circRNAs) are a type of RNA that originates through back-splicing events from linear primary transcripts. CircRNAs display high structural resistance and tissue specificity. Accurate quantification of the circRNA expression level is of vital importance to disease diagnosis. Herein, we construct a label-free fluorescent biosensor for ultrasensitive analysis of circRNAs based on the integration of target-initiated cascade signal amplification strategy with a light-up G-quadruplex. This assay involves only one assistant probe that targets the circRNA-specific back-splice junction. When circRNA is present, it hybridizes with the assistant probe to initiate the duplex-specific nuclease (DSN)-catalyzed cyclic cleavage reaction, producing abundant triggers with 3′OH termini. Then, terminal deoxynucleotidyl transferase (TdT) catalyzes the addition of dGTP and dATP at the 3′-OH termini of the resultant triggers to obtain abundant long G-rich DNA sequences that can form efficient G-quadruplex products. The addition of Thioflavin T (ThT) can light up G-quadruplex, generating an enhanced fluorescence. This assay may be performed isothermally without the involvement of any nucleic acid templates, exogenous primers, and specific labeled probes. Importantly, this biosensor can discriminate target circRNA from one-base mismatched circRNA and exhibits good performance in human serum. Moreover, it can accurately detect circRNA in cancer cells at a single-cell level and even differentiate the circRNA levels in the tissues of healthy persons and nonsmall cell lung cancer (NSCLC) patients, with promising applications in circRNA-related cancer diagnosis and therapeutics.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Target-Initiated Cascade Signal Amplification Lights up a G-Quadruplex for a Label-Free Detection of Circular Ribonucleic Acids

Wei

et al. 2022

Anal. Chem.

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“…It can distinguish circRNAs from corresponding linear transcripts and detect target circRNA as low as 1.1 fM (Liu et al 2020). On top of that, RT-RCA can also be combined with thermostatic netlike hybridization chain reaction (HCR) to detect genes containing repeated sequences, such as telomere DNA, centromeric DNA and ribosomal DNA (rDNA) (Dong et al 2021).…”

Section: Rolling Circle Amplificationmentioning

confidence: 99%

CircRNAs and their regulatory roles in cancers

Mei

Zheng

Yan

et al. 2021

Mol Med

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Circular RNAs (circRNAs), a novel type of non-coding RNAs (ncRNAs), have a covalently closed circular structure resulting from pre-mRNA back splicing via spliceosome and ribozymes. They can be classified differently in accordance with different criteria. As circRNAs are abundant, conserved, and stable, they can be used as diagnostic markers in various diseases and targets to develop new therapies. There are various functions of circRNAs, including sponge for miR/proteins, role of scaffolds, templates for translation, and regulators of mRNA translation and stability. Without m7G cap and poly-A tail, circRNAs can still be degraded in several ways, including RNase L, Ago-dependent, and Ago-independent degradation. Increasing evidence indicates that circRNAs can be modified by N-6 methylation (m6A) in many aspects such as biogenesis, nuclear export, translation, and degradation. In addition, they have been proved to play a regulatory role in the progression of various cancers. Recently, methods of detecting circRNAs with high sensitivity and specificity have also been reported. This review presents a detailed overview of circRNAs regarding biogenesis, biomarker, functions, degradation, and dynamic modification as well as their regulatory roles in various cancers. It’s particularly summarized in detail in the biogenesis of circRNAs, regulation of circRNAs by m6A modification and mechanisms by which circRNAs affect tumor progression respectively. Moreover, existing circRNA detection methods and their characteristics are also mentioned.

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“…The HCR technique is limited by the hybridization kinetics of complementary strands and toehold exchange. It needs a high concentration of DNA or long sticky ends to retain the fast dynamic reaction [23][24][25]. In addition, due to the restriction of enzyme sensitivity, such as temperature and pH, the reaction conditions of ESDA and RCA must be strictly controlled [26,27].…”

Section: Introductionmentioning

confidence: 99%

Chaperone Copolymer Assisted G-Quadruplex-Based Signal Amplification Assay for Highly Sensitive Detection of VEGF

et al. 2022

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Vascular endothelial growth factor (VEGF) is a critical biomarker in the angiogenesis of several cancers. Nowadays, novel approaches to rapid, sensitive, and reliable VEGF detection are urgently required for early cancer diagnosis. Cationic comb-type copolymer, poly(L-lysine)-graft-dextran (PLL-g-Dex) accelerates DNA hybridization and chain exchange reaction while stabilizing the DNA assembly structure. In this work, we examined the chaperone activity of PLL-g-Dex to assist G-quadruplex-based fluorescent DNA biosensors for sensitive detection of VEGF. This convenient and effective strategy is based on chitosan hydrogel, c-myc, Thioflavin T (ThT), VEGF aptamer, and its partially complementary strand. The results show that chaperone copolymer PLL-g-Dex significantly promotes the accumulation of G-quadruplex and assembles into G-wires, allowing an effective signal amplification. Using this method, the detection limit of VEGF was as low as 23 pM, better than many previous works on aptamer-based VEGF detection. This chaperone copolymer-assisted signal amplification strategy has potential applications in the highly sensitive detection of target proteins, even including viruses.

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Specific and sensitive detection of CircRNA based on netlike hybridization chain reaction

Cited by 36 publications

References 44 publications

Target-Initiated Cascade Signal Amplification Lights up a G-Quadruplex for a Label-Free Detection of Circular Ribonucleic Acids

Target-Initiated Cascade Signal Amplification Lights up a G-Quadruplex for a Label-Free Detection of Circular Ribonucleic Acids

CircRNAs and their regulatory roles in cancers

Chaperone Copolymer Assisted G-Quadruplex-Based Signal Amplification Assay for Highly Sensitive Detection of VEGF

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