Abstract Rolling circle amplification (RCA) is an efficient enzymatic isothermal reaction that using circular probe as a template to generate long tandem single-stranded DNA or RNA products under the initiation of short DNA or RNA primers. As a simplified derivative of natural rolling circle replication which synthesizes copies of circular nucleic acids molecules such as plasmids, RCA amplifies the circular template rapidly without thermal cycling and finds various applications in molecular biology. Compared with other amplification strategies, RCA has many obvious advantages. Firstly, because of the strict complementarity required in ligation of a padlock probe, it endows the RCA reaction with high specificity and can even be utilized to distinguish single base mismatches. Secondly, through the introduction of multiple primers, exponential amplification can be achieved easily and leads to a good sensitivity. Thirdly, RCA products can be customized by manipulating circular templates to generate functional nucleic acids such as aptamer, DNAzymes and restriction enzyme sites. Moreover, the RCA has good biocompatibility and is especially suitable for in situ detection. Therefore, RCA has attracted considerable attention as an efficient and potential tool for highly sensitive detection of biomarkers. Herein, we comprehensively introduce the fundamental principles of RCA technology, summarize it from three aspects including initiation mode, amplification mode and signal output mode, and discuss the recent application of RCA-based biosensor in this review.
Last. Mats Nilsson(Science for Life Laboratory)H-Index: 46
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Visualization of the transcriptome in situ has proven to be a valuable tool in exploring single-cell RNA-sequencing data, providing an additional spatial dimension to investigate multiplexed gene expression, cell types, disease architecture or even data driven discoveries. In situ sequencing (ISS) method based on padlock probes and rolling circle amplification has been used to spatially resolve gene transcripts in tissue sections of various origins. Here, we describe the next iteration of ISS, H...
Abstract We present a facile method based on the coffee ring effect that can rapidly detect antibiotic-resistant bacteria, as an affordable genetic testing platform. When a colloidal solution of particles is dropped onto a substrate surface, an outward capillary flow upon evaporation induces the migration of the particles to the periphery of the droplet, forming a characteristic ring pattern. Herein, we utilize capture DNA microbeads which in the presence of target nucleic acid, form suppressed ...
Last. Yongxi Zhao(Xi'an Jiaotong University)H-Index: 24
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Monitoring RNA synthesis and spatial distribution can help to understand its role in physiology and diseases. However, visualiz-ing newly synthesized RNA in single cells remains a great challenge. ...
Accurate identification of exosomes plays an essential role in facilitating disease diagnosis and therapies. Herein, we proposed an Aptamer-cholesterol-mediated Proximity Ligation Assay (AcmPLA) for accurate identification of exosomes in a dual-probe strategy, one aptamer probe for recognition of exosomal innate surface protein CD63 and another cholesterol probe for biolipid layer targeting. By integrating a proximity ligation of probes bound with exosomal biomarkers for specific recognition and...
Abstract Host proteins incorporated into virus particles have been reported to contribute to infectivity and tissue-tropism. This incorporation of host proteins is expected to be variable among viral particles, however, protein analysis at single-virus levels has been challenging. We have developed a method to detect host proteins incorporated on the surface of virions using the in situ proximity ligation assay (isPLA) with rolling circle amplification (RCA), employing oligonucleotide-conjugated...
Last. Dongjiao Li(LYU: Linyi University)H-Index: 1
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Abstract An amplified and highly sensitive hyper-dendritic rolling circle amplification (HRCA) is presented herein. This HRCA system consists of only one circular nucleic acid template that contains three functional areas. One part is an identification sequence of the analyte RNA. The other part is a sequence identical to the molecular beacon modified with a fluorophore/quencher pair. The third part is a sequence identical to the loop region of the coded hairpin structure. These three sequences ...
Abstract Avian influenza virus (AIVs), hosted in poultry, are the pathogens of many poultry diseases and human infections, which bring huge losses to the poultry breeding industry and huge panic to society. Therefore, it is of great significance to establish accurate and sensitive detection methods for AIVs. In this work, a dual-mode detection method based on immuno-rolling circle amplification (immuno-RCA) and quantum dots (QDs) labeling for inductively coupled plasma mass spectrometry (ICP-MS)...
Last. Yongxi Zhao(Xi'an Jiaotong University)H-Index: 24
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5-Hydroxymethyluracil (5hmU) is found in the genomes of a diverse range of organisms as another kind of 5-hydroxymethylpyrimidines except 5-hydroxymethylcytosine (5hmC). The biological function of 5hmU have not been well explored due to lacking both specific 5hmU recognition and single-cell analysis methods. Here we report differentiated visualization of single-cell 5hmU and 5hmC with microfluidic hydrogel encoding (sc5hmU/5hmC-microgel). Single cells and their genomic DNA after cell lysis can b...
Microfluidic techniques are widely used for high-throughput quantification and discrete analysis of micron-scale objects like cells, but are difficult to apply to molecular-scale targets. Instead, single-molecule methods primarily rely on low-throughput imaging of immobilized molecules using high-resolution microscopy. Here we report that commercial-grade flow cytometers can detect single nucleic acid targets following enzymatic extension and dense labeling with multiple distinct fluorophores. W...
Last. Yun Xiang(SWU: Southwest University)H-Index: 51
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MicroRNAs (miRNAs) are useful biomarkers for the diagnosis of a variety of cancers. However, it is of a major challenge to detect miRNAs, considering their high sequence similarity, low concentration and small size nature. With the establishment of an efficient rolling circle amplification (RCA) molecular network by target-driven polymerization/nicking reactions, we present here an exponential amplification strategy for detecting miRNA in a label-free way with ultrahigh sensitivity. The target m...
Abstract null null The importance of user-friendly, inexpensive, sensitive, and selective detection of viruses has been highlighted again due to the recent Coronavirus disease 2019 (COVID-19) pandemic. Among the analytical tools, paper-based devices (PADs) have become a leading alternative for point-of-care (POC) testing. In this review, we discuss the recent development strategies and applications in nucleic acid-based, antibody/antigen-based and other affinity-based PADs using optical and elec...
At present, carcinoembryonic antigen (CEA) is a commonly used biomarker to diagnose cancers. Accurate and specific detection of CEA in clinical samples is of great significance for the diagnosis, treatment and prognosis of related cancers. In this research, we developed an enzyme-free electrochemical apatmer biosensor for highly sensitive detection of CEA based on tetrahedral DNA nanostructures (TDNs) and target-assisted catalytic hairpin assembly (CHA). A long single strand (Trigger), five olig...
#1Lei Wang(CAU: China Agricultural University)H-Index: 5
Last. Jianhan Lin(CAU: China Agricultural University)H-Index: 19
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Bioaerosols containing pathogenic microorganisms have posed a great threat to human and animal health. Effective monitoring of bioaerosols containing pathogenic viruses and bacteria is of great significance to prevent and control infectious diseases. This Feature summarizes recent advances on bioaerosol collection and detection based on microfluidic chips. Besides, the challenges and trends for bioaerosol collection and detection were also discussed.
#2Guangchun Xiao(Qilu University of Technology)H-Index: 9
Last. Xu Chonghai(Qilu University of Technology)H-Index: 1
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Colorectal cancer (CRC) is associated with one of the highest rates of mortality among cancers worldwide. The early detection and management of CRC is imperative. Biomarkers play an important role in CRC screening tests, CRC treatment, and prognosis and clinical management; thus rapid and sensitive detection of biomarkers is helpful for early detection of CRC. In recent years, electrochemical biosensors for detecting CRC biomarkers have been widely investigated. In this review, different electro...