Development of lateral flow assay combined with recombinase polymerase amplification for highly sensitive detection of Dickeya solani

Иванов, А. В.; Safenkova, Irina V.; Drenova, Natalia V.; Zherdev, Anatoly V.; Dzantiev, Boris B.

doi:10.1016/j.mcp.2020.101622

Cited by 21 publications

(12 citation statements)

References 32 publications

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“…Therefore, RPA primers AMV F3/AMV R3 ( Table 1 ) were designed by extension of AMV F1/AMV R1 primers to make them a more appropriate length for RPA. Such primer extension can improve RPA sensitivity [ 10 , 37 ]. Additionally, a new pair for the AMV CP gene, AMV F4/AMV R4, was designed ( Table 1 ).…”

Section: Resultsmentioning

confidence: 99%

“…The easiest way to obtain the labeled amplicon is by using forward and reverse RPA primers labeled at the 5′ ends with different tags (the scheme is presented in Figure 1 A). This approach is actively applied for the RPA-LFT detection of pathogens and species identification [ 8 , 9 , 10 , 11 ]. However, the probability of the cross-dimerization of the labeled RPA primers is high due to the typical RPA primers being rather long (28–35 nt), and there are no stages with the temperature denaturation of DNA.…”

Section: Introductionmentioning

confidence: 99%

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Recombinase Polymerase Amplification Assay with and without Nuclease-Dependent-Labeled Oligonucleotide Probe

Иванов

Safenkova

Zherdev

et al. 2021

IJMS

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The combination of recombinase polymerase amplification (RPA) and lateral flow test (LFT) is a strong diagnostic tool for rapid pathogen detection in resource-limited conditions. Here, we compared two methods generating labeled RPA amplicons following their detection by LFT: (1) the basic one with primers modified with different tags at the terminals and (2) the nuclease-dependent one with the primers and labeled oligonucleotide probe for nuclease digestion that was recommended for the high specificity of the assay. Using both methods, we developed an RPA-LFT assay for the detection of worldwide distributed phytopathogen—alfalfa mosaic virus (AMV). A forward primer modified with fluorescein and a reverse primer with biotin and fluorescein-labeled oligonucleotide probe were designed and verified by RPA. Both labeling approaches and their related assays were characterized using the in vitro-transcribed mRNA of AMV and reverse transcription reaction. The results demonstrated that the RPA-LFT assay based on primers-labeling detected 103 copies of RNA in reaction during 30 min and had a half-maximal binding concentration 22 times lower than probe-dependent RPA-LFT. The developed RPA-LFT was successfully applied for the detection of AMV-infected plants. The results can be the main reason for choosing simple labeling with primers for RPA-LFT for the detection of other pathogens.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Recombinase Polymerase Amplification Assay with and without Nuclease-Dependent-Labeled Oligonucleotide Probe

Иванов

Safenkova

Zherdev

et al. 2021

IJMS

Self Cite

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“…RPA was performed with FAM-and biotin-labelled primers that were picked after screening. This simplification has previously been applied in other RPA papers [212][213][214]223]. Of course, an additional probe in RPA should enhance specificity and decrease background on the test dipstick.…”

Section: Rpa-based Testsmentioning

confidence: 98%

“…First, all these tests were performed with the following commercial RPA kits: TwistDx nfo [27,84,[208][209][210][211], TwistDx Basic [212][213][214][215], TwistDx Exo [51], and Amplifier (Agdia) [27,92,[216][217][218]. The main criterion for the selection of a gene as a target for RPA was maximal specificity for the species of interest.…”

Section: Rpa-based Testsmentioning

confidence: 99%

“…Some tests utilize anti-digoxygenin conjugate with carbon NPs in the test zone [211]. The commercial LFA kits were purchased from Millenia [51,[208][209][210], PCRD nucleic acid detector [84,211,222], BioUSTAR [26], and Agdia [27,92,[216][217][218] or assembled and optimized from different membranes (homemade test strips) [212][213][214]. RPA contains a large amount of DTT that induces aggregation of some GNP conjugates, such as streptavidin-GNP [87].…”

Section: Rpa-based Testsmentioning

confidence: 99%

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The Potential Use of Isothermal Amplification Assays for In-Field Diagnostics of Plant Pathogens

Иванов

Safenkova

Zherdev

et al. 2021

Plants

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Rapid, sensitive, and timely diagnostics are essential for protecting plants from pathogens. Commonly, PCR techniques are used in laboratories for highly sensitive detection of DNA/RNA from viral, viroid, bacterial, and fungal pathogens of plants. However, using PCR-based methods for in-field diagnostics is a challenge and sometimes nearly impossible. With the advent of isothermal amplification methods, which provide amplification of nucleic acids at a certain temperature and do not require thermocyclic equipment, going beyond the laboratory has become a reality for molecular diagnostics. The amplification stage ceases to be limited by time and instruments. Challenges to solve involve finding suitable approaches for rapid and user-friendly plant preparation and detection of amplicons after amplification. Here, we summarize approaches for in-field diagnostics of phytopathogens based on different types of isothermal amplification and discuss their advantages and disadvantages. In this review, we consider a combination of isothermal amplification methods with extraction and detection methods compatible with in-field phytodiagnostics. Molecular diagnostics in out-of-lab conditions are of particular importance for protecting against viral, bacterial, and fungal phytopathogens in order to quickly prevent and control the spread of disease. We believe that the development of rapid, sensitive, and equipment-free nucleic acid detection methods is the future of phytodiagnostics, and its benefits are already visible.

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Novel multiplex recombinase polymerase amplification assay for simultaneous detection and serotyping of Streptococcus parauberis

Choi,

Kim,

Kim

2024

Aquacult Int

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Development of lateral flow assay combined with recombinase polymerase amplification for highly sensitive detection of Dickeya solani

Cited by 21 publications

References 32 publications

Recombinase Polymerase Amplification Assay with and without Nuclease-Dependent-Labeled Oligonucleotide Probe

Recombinase Polymerase Amplification Assay with and without Nuclease-Dependent-Labeled Oligonucleotide Probe

The Potential Use of Isothermal Amplification Assays for In-Field Diagnostics of Plant Pathogens

Novel multiplex recombinase polymerase amplification assay for simultaneous detection and serotyping of Streptococcus parauberis

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