RPA-SYBR Green I based instrument-free visual detection for pathogenic Yersinia enterocolitica in meat

Zheng, Yu; Hu, Pan; Ren, Hong-Lin; Wang, Han; Cao, Qi; Zhao, Qiang; Li, Hanxiao; Zhang, Hailing; Liu, Zhanxu; Li, Yansong; Liu, Zengshan; Lu, Shiying

doi:10.1016/j.ab.2021.114157

Cited by 20 publications

(8 citation statements)

References 20 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The real-time RPA sensitivity experiments can detect a minimum concentration of 10 −4 ng/μL, which is almost identical to RT-PCR [ 4 ] and LAMP [ 15 ] sensitivity. It is 10 times more sensitive compared to RPA-SYBR Green I [ 32 ] and normal PCR. When the amount of DNA is 35 ng, it can show significant amplification within 3 min with an obvious fluorescence signal, which can rapidly detect Y. enterocolitica .…”

Section: Discussionmentioning

confidence: 99%

Establishment of a Real-Time Recombinase Polymerase Amplification for Rapid Detection of Pathogenic Yersinia enterocolitica

Zhang

Zhao

et al. 2023

Pathogens

View full text Add to dashboard Cite

Yersinia enterocolitica is a zoonotic proto-microbe that is widespread throughout the world, causes self-limiting diseases in humans or animals and even leads to sepsis and death in patients with severe cases. In this study, a real-time recombinase polymerase amplification (RPA) assay for pathogenic Y. enterocolitica was established based on the ail gene. The results showed that the RPA detection for Y. enterocolitica could be completed within 20 min at an isothermal temperature of 38 °C by optimizing the conditions in the primers and Exo probe. Moreover, the sensitivity of the current RT-RPA was 10−4 ng/μL, and the study found that the assay was negative in the application of the genomic DNA of other pathogens. These suggest the establishment of a rapid and sensitive real-time RPA method for the detection of pathogenic Y. enterocolitica, which can provide new understandings for the early diagnosis of the pathogens.

show abstract

Section: Discussionmentioning

confidence: 99%

Establishment of a Real-Time Recombinase Polymerase Amplification for Rapid Detection of Pathogenic Yersinia enterocolitica

Zhang

Zhao

et al. 2023

Pathogens

View full text Add to dashboard Cite

show abstract

“…Among them, RPA has been widely used for nucleic acid detection because of its high efficiency and easily achievable reaction temperature (37 -42 ℃) [15]. Recently, strategies such as RPA-AGE (RPA combined with agarose gel electrophoresis) [16], RT-RPA (real-time fluorescent RPA) [15,17], RPA-SYBR Green I (RPA combined with DNA fluorescent dye) [18,19], RPA-LFD (RPA combined with lateral flow immunochromatography) [20,21] and RPA-Cas12a-FS (RPA combined with Cas12a cutting) [22] have been established and implemented in the field of rapid detection. Among these methods, the test strip method is the most suitable for on-site testing for its full independence of testing equipment.…”

Section: Introductionmentioning

confidence: 99%

A dual-RPA based lateral flow strip for sensitive, on-site detection of CP4- EPSPS and Cry1Ab/ Ac genes in genetically modified crops

Wang¹,

Wang²,

Hu³

et al. 2024

Food Science and Human Wellness

View full text Add to dashboard Cite

Traditional transgenic detection methods require high test conditions and struggle to be both sensitive and efficient. In this study, a one-tube dual RPA reaction system for CP4-EPSPS and Cry1Ab/Ac was proposed and combined with a lateral flow immunochromatographic assay, named "Dual-RPA-LFD", to visualize the dual detection of GM crops. In which, the herbicide tolerance gene CP4-EPSPS and the insect resistance gene Cry1Ab/Ac were selected as targets taking into account the current status of the most widespread application of insect resistance and herbicide tolerance traits and their stacked traits. Gradient diluted plasmids, transgenic standards, and actual samples were used as templates to conduct sensitivity, specificity, and practicality assays, respectively. The constructed method achieved the visual detection of plasmid at levels as low as 100 copies, demonstrating its high sensitivity. In addition, good applicability to transgenic samples was observed, with no cross-interference between two test lines and no influence from other genes. In conclusion, this strategy achieved the expected purpose of simultaneous detection of the two popular targets in GM crops within 20 minutes at 37 ℃ in a rapid, equipment-free field manner, providing a new alternative for rapid screening for transgenic assays in the field.

show abstract

“…RPA simplifies assay design and reduces the risk of primer interactions or cross-reactivity by employing a fewer number of primers in comparison to LAMP . DNA can be amplified in 20 min at 37 to 42 °C with minimal effort and at room temperature using RPA. , Due to these advantages, RPA has been used in a wide variety of meat adulteration detection methods. – Nonetheless, the specificity of these approaches can be enhanced so that they can be applied in a wider variety of situations.…”

Section: Introductionmentioning

confidence: 99%

Amplification-Based CRISPR/Cas12a Biosensor Targeting the COX1 Gene for Specific Detection of Porcine DNA

Janudin,

Kurup,

Chee

et al. 2023

ACS Omega

View full text Add to dashboard Cite

We propose a CRISPR/Cas12a-mediated recombinase polymerase amplification (RPA) detection method that combines RPA with Cas12a cleavage for the detection of halal food adulteration, which is of global concern, particularly for Muslim consumers. We optimized the reagent concentrations for the Cas12a cleavage steps and designed and screened gRNA targeting a conserved area of the mitochondrial cytochrome C oxidase subunit I (COX1) gene. This procedure successfully detected the presence of porcine components as low as 5 pg/μL in the linear range of 5−1000 pg/μL. The assay's detection limit was 500 times lower than CRISPR-based approaches that exclude a preamplification step, allowing the detection of trace porcine DNA in food samples. The assay additionally showed no cross-reaction with nontarget species. Therefore, this detection platform shows tremendous potential as a method for the quick, sensitive, and specific detection of porcine-derived components.

show abstract

RPA-SYBR Green I based instrument-free visual detection for pathogenic Yersinia enterocolitica in meat

Cited by 20 publications

References 20 publications

Establishment of a Real-Time Recombinase Polymerase Amplification for Rapid Detection of Pathogenic Yersinia enterocolitica

Establishment of a Real-Time Recombinase Polymerase Amplification for Rapid Detection of Pathogenic Yersinia enterocolitica

A dual-RPA based lateral flow strip for sensitive, on-site detection of CP4- EPSPS and Cry1Ab/ Ac genes in genetically modified crops

Amplification-Based CRISPR/Cas12a Biosensor Targeting the COX1 Gene for Specific Detection of Porcine DNA

Contact Info

Product

Resources

About