Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4
Objective To establish a fluorescent reverse transcription-recombinase polymerase amplification (RT-RPA) assay for norovirus (NoV) type GⅡ.4. Methods The NoV GⅡ.4 gene sequence was obtained from NCBI, and the primers and probes were designed by selecting the conserved regions after sequence alignmen...
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Editorial Office of Journal of Precision Medicine
2025-04-01
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| Series: | 精准医学杂志 |
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| Online Access: | https://jpmed.qdu.edu.cn/fileup/2096-529X/PDF/1745982310804-159985325.pdf |
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| author | ZHENG Zhikun, XU Hefei, DING Wei, ZHANG Juan, DU Jiansen, HOU Lin, ZHANG Jin |
| author_facet | ZHENG Zhikun, XU Hefei, DING Wei, ZHANG Juan, DU Jiansen, HOU Lin, ZHANG Jin |
| author_sort | ZHENG Zhikun, XU Hefei, DING Wei, ZHANG Juan, DU Jiansen, HOU Lin, ZHANG Jin |
| collection | DOAJ |
| description | Objective To establish a fluorescent reverse transcription-recombinase polymerase amplification (RT-RPA) assay for norovirus (NoV) type GⅡ.4. Methods The NoV GⅡ.4 gene sequence was obtained from NCBI, and the primers and probes were designed by selecting the conserved regions after sequence alignment. With NoV GⅡ.4 standards as the detection object, the temperature, primer concentration, and probe concentration of the fluorescent RT-RPA reaction system were optimized to establish a fluorescent RT-RPA assay for NoV GⅡ.4. This method was assessed in terms of sensitivity, specificity, and clinical validation by detecting different concentrations of NoV GⅡ.4 standards and the nucleic acids of NoV GⅠ, other subtypes of NoV GⅡ (NoV GⅡ.2, NoV GⅡ.3, NoV GⅡ.7, NoV GⅡ.8, and NoV GⅡ.17), rotavirus, adenovirus, parechovirus, astrovirus, and sapovirus. Results The fluorescent RT-RPA assay had an optimal reaction temperature of 41 ℃, an optimal primer concentration of 630 nmol/L, and an optimal probe concentration of 240 nmol/L for detecting NoV GⅡ.4. And the limit of detection was 1×101 copies/μL for NoV GⅡ.4, the other nucleic acids did not produce the fluorescent signal that could be detected by the instrument. Conclusion This study establishes a fluorescent RT-RPA assay for NoV GⅡ.4, which is characterized by high sensitivity, good specificity, and rapid reaction speed, and it is expected to provide a new method for the rapid detection of NoV GⅡ.4 at ports. |
| format | Article |
| id | doaj-art-88d099261e414277a9d9fa04bdbcc000 |
| institution | OA Journals |
| issn | 2096-529X |
| language | zho |
| publishDate | 2025-04-01 |
| publisher | Editorial Office of Journal of Precision Medicine |
| record_format | Article |
| series | 精准医学杂志 |
| spelling | doaj-art-88d099261e414277a9d9fa04bdbcc0002025-08-20T02:28:47ZzhoEditorial Office of Journal of Precision Medicine精准医学杂志2096-529X2025-04-0140212713010.13362/j.jpmed.202540038Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4ZHENG Zhikun, XU Hefei, DING Wei, ZHANG Juan, DU Jiansen, HOU Lin, ZHANG Jin0Department of Biochemistry and Molecular Biology, College of Basic Medical Sciences, Qingdao University, Qingdao 266071, ChinaObjective To establish a fluorescent reverse transcription-recombinase polymerase amplification (RT-RPA) assay for norovirus (NoV) type GⅡ.4. Methods The NoV GⅡ.4 gene sequence was obtained from NCBI, and the primers and probes were designed by selecting the conserved regions after sequence alignment. With NoV GⅡ.4 standards as the detection object, the temperature, primer concentration, and probe concentration of the fluorescent RT-RPA reaction system were optimized to establish a fluorescent RT-RPA assay for NoV GⅡ.4. This method was assessed in terms of sensitivity, specificity, and clinical validation by detecting different concentrations of NoV GⅡ.4 standards and the nucleic acids of NoV GⅠ, other subtypes of NoV GⅡ (NoV GⅡ.2, NoV GⅡ.3, NoV GⅡ.7, NoV GⅡ.8, and NoV GⅡ.17), rotavirus, adenovirus, parechovirus, astrovirus, and sapovirus. Results The fluorescent RT-RPA assay had an optimal reaction temperature of 41 ℃, an optimal primer concentration of 630 nmol/L, and an optimal probe concentration of 240 nmol/L for detecting NoV GⅡ.4. And the limit of detection was 1×101 copies/μL for NoV GⅡ.4, the other nucleic acids did not produce the fluorescent signal that could be detected by the instrument. Conclusion This study establishes a fluorescent RT-RPA assay for NoV GⅡ.4, which is characterized by high sensitivity, good specificity, and rapid reaction speed, and it is expected to provide a new method for the rapid detection of NoV GⅡ.4 at ports.https://jpmed.qdu.edu.cn/fileup/2096-529X/PDF/1745982310804-159985325.pdfnorovirus|recombinase polymerase amplification|nucleic acid amplification techniques|sensitivity and specificity |
| spellingShingle | ZHENG Zhikun, XU Hefei, DING Wei, ZHANG Juan, DU Jiansen, HOU Lin, ZHANG Jin Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4 精准医学杂志 norovirus|recombinase polymerase amplification|nucleic acid amplification techniques|sensitivity and specificity |
| title | Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4 |
| title_full | Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4 |
| title_fullStr | Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4 |
| title_full_unstemmed | Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4 |
| title_short | Establishment of a fluorescent reverse transcription-recombinase polymerase amplification assay for norovirus type GⅡ.4 |
| title_sort | establishment of a fluorescent reverse transcription recombinase polymerase amplification assay for norovirus type gii 4 |
| topic | norovirus|recombinase polymerase amplification|nucleic acid amplification techniques|sensitivity and specificity |
| url | https://jpmed.qdu.edu.cn/fileup/2096-529X/PDF/1745982310804-159985325.pdf |
| work_keys_str_mv | AT zhengzhikunxuhefeidingweizhangjuandujiansenhoulinzhangjin establishmentofafluorescentreversetranscriptionrecombinasepolymeraseamplificationassayfornorovirustypegii4 |