RPB0322

Pathogen Description

Target Pathogen Pathogen Name NCBI Taxonomy ID Order Family Genus Species Pathogen type
Influenza A virus (H9N2) Influenza A virus (A\chicken\Yunnan\Baoshan2\2007(H9N2)) 479038 Articulavirales Orthomyxoviridae Alphainfluenzavirus Influenza A virus Virus

Primer Description

Primer Name Sequence(5'-3') Length(bp) Primer Final Concentration(μM） GC Content（%） Predicted Melting Temperature(℃） Molecular Weight(g/moles) Positions in GenBank accession number
RPA-F4 TAATACGACTCACTATAGGGCGCAGATCTCGAGGCTCTC 39 10 μM 51.28 67.38 11951.81 \
RPA-R4 GCCTTGTCCATGTTGTTTGGGTCTCCATTCCC 32 10 μM 53.13 66.59 9715.32 \

Gene Description

Target Gene GenBank ID
M gene MK447276.1

Assay Description

Application Assay Primer Designing Software Reaction Time(min) Assay Temperature(℃) Readout System(s) Limit of Detection(LoD) Sensitivity(%) Specificity(%)
high reliability, sensitivity, and specificity, with promising potential for rapidly detecting H9N2 RPA-CRISPR\Cas13a Lasergene 12 Core Suite 40 min 37 °C CRISPR-Cas13a \ 10 copies/μL \

Publication Description

Year of Publication Title Author(s) Journal PMID DOI
2025 H9N2 avian influenza virus diagnostics utilizing specific high-sensitivity enzymatic molecular system termed RPA-based CRISPR-Cas13a Dalin He,Saisai Zhao,Fangfang Wang,Bingrong Wu,Feng Wei,Yubo Zhao,Xinhui Wei,Hui Ren,Meijuan Zhang,Yaru Fan,Jiahao Zhang,Shumin Yu,Yi Tang,Youxiang Diao International Journal of Biological Macromolecules 39884612 10.1016/j.ijbiomac.2025.140474

H9N2 avian influenza virus diagnostics utilizing specific high-sensitivity enzymatic molecular system termed RPA-based CRISPR-Cas13a

Author(s):

Dalin He,Saisai Zhao,Fangfang Wang,Bingrong Wu,Feng Wei,Yubo Zhao,Xinhui Wei,Hui Ren,Meijuan Zhang,Yaru Fan,Jiahao Zhang,Shumin Yu,Yi Tang,Youxiang Diao

Journal:

International Journal of Biological Macromolecules

Year:

2025

Abstract:

H9N2 avian influenza virus (AIV), a major pathogen causing respiratory infections in poultry, poses a significant threat to the poultry industry and human health. Early detection and control of H9N2 infections are essential for minimizing economic losses and preventing potential zoonotic transmission. A novel CRISPR-Cas family member called CRISPR-Cas13a comprises the CRISPR RNA (crRNA) and Cas13a nuclease. Through the crRNA-based reprogramming of Cas13a, a platform for sensing RNAs specifically is available. In this study, we developed a RPA-based CRISPR-Cas13a diagnostic method for rapid detection of the H9N2 AIV. The results demonstrated that at a limit of 10 copies/μL and 102 copies/μL could be detected within 50 min, by fluorescence detection and lateral flow strip, respectively, offering a highly sensitive method for H9N2 detection. This method exhibited excellent specificity, distinguishing H9N2 from other pathogens. Furthermore, the RPA-Cas13a-based detection system was tested on clinical samples, showing comparable performance to RT-qPCR. The detection results were visualized using either lateral flow assays or fluorescence, making it a suitable tool for on-site, field-deployable diagnostics. In a word, this RPA-Cas13a diagnostic approach offers high reliability, sensitivity, and specificity, with promising potential for rapidly detecting H9N2 and other viral pathogens in clinical and food safety applications.