Specific detection of topoisomerase i from the malaria causing P. falciparum parasite using isothermal Rolling Circle Amplification

2012 ◽  
Author(s):  
C. Tesauro ◽  
S. Juul ◽  
B. Arno ◽  
C. J. F. Nielsen ◽  
P. Fiorani ◽  
...  
Keyword(s):  
Topoisomerase I ◽  
Rolling Circle Amplification ◽  
Specific Detection ◽  
Rolling Circle ◽  
Falciparum Parasite

Molecular Beacon Enables Combination of Highly Processive and Highly Sensitive Rolling Circle Amplification Readouts for Detection of DNA-Modifying Enzymes

Nano LIFE ◽  
2015 ◽  
Vol 05 (02) ◽  
pp. 1541002 ◽  
Author(s):  
Emil L. Kristoffersen ◽  
Maria Gonzalez ◽  
Magnus Stougaard ◽  
Cinzia Tesauro
Keyword(s):  
Real Time ◽  
Dna Cleavage ◽  
Topoisomerase I ◽  
Drug Response ◽  
Molecular Beacon ◽  
Rolling Circle Amplification ◽  
Dna Topoisomerase ◽  
Rolling Circle ◽  
Topoisomerase Ib ◽  
Highly Sensitive

Here we present an optimized readout format for detection of the circularized products from a DNA-based sensor for measurement of DNA-modifying enzymes including DNA Topoisomerase I. The basic design of the DNA-sensor relies on the use of a substrate that can be circularized by the activity of DNA-modifying enzymes like type IB Topoisomerases and subsequently amplified by a rolling circle amplification (RCA) mechanism. The RCA process can be followed in real-time by the addition of a molecular beacon with a fluorophore/quencher pair. Upon hybridization to the amplified product, the fluorophore/quencher pair is separated, giving rise to a fluorescent signal, measurable in pseudo real-time using a qPCR machine or in a fluorimeter. The RCA products in complex with the molecular beacon can subsequently be moved to microscopic slides and analyzed in a fluorescence microscope. We describe the proof of the principle of this molecular beacon-based method combining the qPCR readout format with the standard Rolling circle Enhanced Enzymatic Assay previously reported. Although the qPCR setup is less sensitive, it allows easy, fast, and high-throughput measurement of enzyme activities. Human Topoisomerase IB (TopIB) is a well-known target for the clinically used anticancer drugs of the camptothecin family. The cytotoxic effect of camptothecins correlates directly with the intracellular TopIB activity affecting reversibly the Topoisomerase/DNA cleavage complexes. Therefore, we envisioned that the presented method may find use for the prediction of cellular drug response and for drug screening to discover novel molecules that specifically inhibit TopIB or other DNA-modifying enzymes.

Protein binding-protected DNA three-way junction-mediated rolling circle amplification for sensitive and specific detection of transcription factors

RSC Advances ◽  
2016 ◽  
Vol 6 (73) ◽  
pp. 68846-68851 ◽  
Author(s):  
Kan Li ◽  
Lei Wang ◽  
Xiaowen Xu ◽  
Ting Gao ◽  
Ping Yan ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Protein Binding ◽  
Rolling Circle Amplification ◽  
Specific Detection ◽  
Rolling Circle

A novel fluorescent strategy for transcription factors assay was developed based on protein binding-protected DNA three-way junction-mediated rolling circle amplification.

DNA-Mediated rolling circle amplification for ultrasensitive detection of thrombin using MALDI-TOF mass spectrometry

2018 ◽  
Vol 54 (82) ◽  
pp. 11546-11549 ◽  
Author(s):  
Zengnan Wu ◽  
Ling Lin ◽  
Mashooq Khan ◽  
Weifei Zhang ◽  
Sifeng Mao ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Rolling Circle Amplification ◽  
Ultrasensitive Detection ◽  
Specific Detection ◽  
Rolling Circle ◽  
Maldi Tof Ms ◽  
Maldi Tof Mass Spectrometry ◽  
Maldi Tof ◽  
Tof Ms

A DNA-mediated rolling circle amplification (RCA) strategy was established for ultrasensitive and specific detection of thrombin via MALDI-TOF MS.

An electrochemiluminescence biosensor for Kras mutations based on locked nucleic acid functionalized DNA walkers and hyperbranched rolling circle amplification

2017 ◽  
Vol 53 (20) ◽  
pp. 2910-2913 ◽  
Author(s):  
Ying Zhang ◽  
Lixu Wang ◽  
Fang Luo ◽  
Bin Qiu ◽  
Longhua Guo ◽  
...  
Keyword(s):  
Nucleic Acid ◽  
Rolling Circle Amplification ◽  
Locked Nucleic Acid ◽  
Specific Detection ◽  
Kras Mutations ◽  
Rolling Circle ◽  
Dna Walkers ◽  
Mutant Genes

Herein, an electrochemiluminescence (ECL) biosensor for ultrasensitive and specific detection of Kras mutant genes has been developed.

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