scholarly journals Ultrafast, one-step, and microwave heating-based synthesis of DNA/RNA-AuNP conjugates

2021 ◽  
Author(s):  
Mengqi Huang ◽  
Erhu Xiong ◽  
Menglu Hu ◽  
Huahua Yue ◽  
Tian Tian ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Rolling Circle Amplification ◽  
Test Strip ◽  
N Gene ◽  
Great Promise ◽  
Rolling Circle ◽  
Spherical Nucleic Acid ◽  
Standing Up ◽  
One Step ◽  
Nucleic Acids Delivery

DNA/RNA-gold nanoparticle (DNA/RNA-AuNP) nanoprobes have been widely employed for nanobiotechnology applications. Here we discovered that both thiolated and non-thiolated DNA/RNA can be efficiently attached to AuNPs to achieve high-stable spherical nucleic acid (SNA) within minutes under a domestic microwave (MW)-assisted heating-dry circumstance. Further studies showed that for non-thiolated DNA/RNA the conjugation is poly (T/U) tag dependent. Spectroscopy, test strip hybridization, and loading counting experiments indicate that low-affinity poly (T/U) tag mediates the formation of a standing-up conformation, which is distributed in the outer layer of such a SNA structure. In further applications study, CRISPR/Cas9-sgRNA (135 bp), RNA from Nucleocapsid (N) gene of SARS-CoV-2 (1279 bp), and rolling circle amplification (RCA) DNA products (over 1000 bp) could be successfully attached on AuNPs, which overcomes the routine methods in long-chain nucleic acid-AuNP conjugation, exhibiting great promise in novel biosensing and nucleic acids delivery strategy. This novel heating-dry strategy has improved the traditional DNA/RNA-AuNP conjugation methods in simplicity, rapidity, cost, and universality.

The analysis of proteins and small molecules based on sterically tunable nucleic acid hyperbranched rolling circle amplification

2017 ◽  
Vol 91 ◽  
pp. 136-142 ◽  
Author(s):  
Hai Shi ◽  
Xiaoxia Mao ◽  
Xiaoxia Chen ◽  
Zihan Wang ◽  
Keming Wang ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Small Molecules ◽  
Rolling Circle Amplification ◽  
Rolling Circle

scholarly journals Rapid electrochemical detection of coronavirus SARS-CoV-2

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Thanyarat Chaibun ◽  
Jiratchaya Puenpa ◽  
Tatchanun Ngamdee ◽  
Nimaradee Boonapatcharoen ◽  
Pornpat Athamanolap ◽  
...  
Keyword(s):  
Electrochemical Biosensor ◽  
Rolling Circle Amplification ◽  
Clinical Samples ◽  
Rolling Circle ◽  
Qrt Pcr ◽  
Reverse Transcription Pcr ◽  
Sandwich Hybridization ◽  
Redox Active ◽  
One Step ◽  
The One

AbstractCoronavirus disease 2019 (COVID-19) is a highly contagious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Diagnosis of COVID-19 depends on quantitative reverse transcription PCR (qRT-PCR), which is time-consuming and requires expensive instrumentation. Here, we report an ultrasensitive electrochemical biosensor based on isothermal rolling circle amplification (RCA) for rapid detection of SARS-CoV-2. The assay involves the hybridization of the RCA amplicons with probes that were functionalized with redox active labels that are detectable by an electrochemical biosensor. The one-step sandwich hybridization assay could detect as low as 1 copy/μL of N and S genes, in less than 2 h. Sensor evaluation with 106 clinical samples, including 41 SARS-CoV-2 positive and 9 samples positive for other respiratory viruses, gave a 100% concordance result with qRT-PCR, with complete correlation between the biosensor current signals and quantitation cycle (Cq) values. In summary, this biosensor could be used as an on-site, real-time diagnostic test for COVID-19.

Mitigating the Non-Specific Amplification in RCA: Assessment of the Role of Ligation and Exonuclease Digestion in Circular DNA Preparation

2021 ◽  
Author(s):  
Vandana Kuttappan Nair ◽  
Chandrika Sharma ◽  
Mrittika Sengupta ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
False Positive ◽  
Rolling Circle Amplification ◽  
Circular Dna ◽  
Rolling Circle ◽  
Specific Amplification ◽  
Exonuclease Digestion ◽  
Sticky End

<b>Layman Summary: </b>Rolling circle amplification (RCA) is a popular and extensively used bioanalytical tool. Like any nucleic acid amplifications, non-specific amplification may occur in it and risk generating false positive readouts. The work described in the manuscript investigates non-specific amplification in RCA as a function of ligation and exonuclease digestion assays during the synthesis of circular DNA. In particular, it investigates and compares the role of three different ligation techniques, namely splint-padlock ligation, cohesive end (sticky end ligation), and self-annealing ligation. In addition, it also probes the role of single exonuclease vs dual exonuclease digestions. We employed real time fluorescence to quantify the effect of these factors. Finally, our work hypothesizes the possible origins of non-specific amplification in RCA.

One-step discrimination of BCR/ABLp210 transcript isoforms directly from RNA extraction with fusion-triggered rolling circle amplification

2019 ◽  
Vol 1067 ◽  
pp. 129-136 ◽  
Author(s):  
Nini Luo ◽  
Qianfeng Xia ◽  
Lutan Zhang ◽  
Yuhong Zhang ◽  
Lizhen Huang ◽  
...  
Keyword(s):  
Rolling Circle Amplification ◽  
Rna Extraction ◽  
Transcript Isoforms ◽  
Rolling Circle ◽  
One Step

Nucleic acid sensing by regenerable surface-associated isothermal rolling circle amplification

2007 ◽  
Vol 22 (7) ◽  
pp. 1236-1244 ◽  
Author(s):  
Erik L. McCarthy ◽  
Lee E. Bickerstaff ◽  
Mauricio Pereira da Cunha ◽  
Paul J. Millard
Keyword(s):  
Nucleic Acid ◽  
Rolling Circle Amplification ◽  
Rolling Circle

scholarly journals Rolling Circle Amplification as an Efficient Analytical Tool for Rapid Detection of Contaminants in Aqueous Environments

Biosensors ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 352
Author(s):  
Kuankuan Zhang ◽  
Hua Zhang ◽  
Haorui Cao ◽  
Yu Jiang ◽  
Kang Mao ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Rolling Circle Amplification ◽  
Environmental Pollutants ◽  
Monitoring Method ◽  
Rolling Circle ◽  
Advantages And Disadvantages ◽  
Contaminant Monitoring ◽  
Aqueous Environments ◽  
Global Concern ◽  
Circular Template

Environmental contaminants are a global concern, and an effective strategy for remediation is to develop a rapid, on-site, and affordable monitoring method. However, this remains challenging, especially with regard to the detection of various contaminants in complex water environments. The application of molecular methods has recently attracted increasing attention; for example, rolling circle amplification (RCA) is an isothermal enzymatic process in which a short nucleic acid primer is amplified to form a long single-stranded nucleic acid using a circular template and special nucleic acid polymerases. Furthermore, this approach can be further engineered into a device for point-of-need monitoring of environmental pollutants. In this paper, we describe the fundamental principles of RCA and the advantages and disadvantages of RCA assays. Then, we discuss the recently developed RCA-based tools for environmental analysis to determine various targets, including heavy metals, organic small molecules, nucleic acids, peptides, proteins, and even microorganisms in aqueous environments. Finally, we summarize the challenges and outline strategies for the advancement of this technique for application in contaminant monitoring.

Mitigating the Non-Specific Amplification in RCA: Assessment of the Role of Ligation and Exonuclease Digestion in Circular DNA Preparation

2021 ◽  
Author(s):  
Vandana Kuttappan Nair ◽  
Chandrika Sharma ◽  
Mrittika Sengupta ◽  
Souradyuti Ghosh
Keyword(s):  
Nucleic Acid ◽  
Real Time ◽  
False Positive ◽  
Rolling Circle Amplification ◽  
Circular Dna ◽  
Rolling Circle ◽  
Specific Amplification ◽  
Exonuclease Digestion ◽  
Sticky End

<b>Layman Summary: </b>Rolling circle amplification (RCA) is a popular and extensively used bioanalytical tool. Like any nucleic acid amplifications, non-specific amplification may occur in it and risk generating false positive readouts. The work described in the manuscript investigates non-specific amplification in RCA as a function of ligation and exonuclease digestion assays during the synthesis of circular DNA. In particular, it investigates and compares the role of three different ligation techniques, namely splint-padlock ligation, cohesive end (sticky end ligation), and self-annealing ligation. In addition, it also probes the role of single exonuclease vs dual exonuclease digestions. We employed real time fluorescence to quantify the effect of these factors. Finally, our work hypothesizes the possible origins of non-specific amplification in RCA.

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