{"title":"基于CRISPR/Cas12a和重组酶聚合酶扩增技术的鸭圆环病毒快速现场核酸检测","authors":"Qi-Zhang Liang, Wei Chen, Rong-Chang Liu, Qiu-Ling Fu, Guang-Hua Fu, Long-Fei Cheng, Hong-Mei Chen, Nan-Song Jiang, Ting Zhu, Yu Huang","doi":"10.1186/s12985-024-02577-7","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Duck circovirus (DuCV) infections commonly induce immunosuppression and secondary infections in ducks, resulting in significant economic losses in the duck breeding industry. Currently, effective vaccines and treatments for DuCV have been lacking. Therefore, rapid, specific, and sensitive detection methods are crucial for preventing and controlling DuCV.</p><p><strong>Methods: </strong>A lateral flow strip (LFS) detection method was developed using recombinase polymerase amplification (RPA) and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 12a (Cas12a). The RPA-CRISPR/Cas12a-LFS targeted the DuCV replication protein (Rep) and was operated at 37 ℃ and allowed for visual interpretation without requiring sophisticated equipment.</p><p><strong>Results: </strong>The results revealed that the reaction time of RPA-CRISPR/Cas12a-LFS is only 45 min. This method achieved a low detection limit of 2.6 gene copies. Importantly, this method demonstrated high specificity and no cross-reactivity with six other avian viruses. In a study involving 97 waterfowl samples, the Rep RPA-CRISPR/Cas12a-LFS showed 100% consistency and agreement with real-time quantitative polymerase chain reaction.</p><p><strong>Conclusion: </strong>These findings underscored the potential of this user-friendly, rapid, sensitive, and accurate detection method for on-site DuCV detection.</p>","PeriodicalId":23616,"journal":{"name":"Virology Journal","volume":"21 1","pages":"322"},"PeriodicalIF":4.0000,"publicationDate":"2024-12-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11661005/pdf/","citationCount":"0","resultStr":"{\"title\":\"CRISPR/Cas12a and recombinase polymerase amplification-based rapid on-site nucleic acid detection of duck circovirus.\",\"authors\":\"Qi-Zhang Liang, Wei Chen, Rong-Chang Liu, Qiu-Ling Fu, Guang-Hua Fu, Long-Fei Cheng, Hong-Mei Chen, Nan-Song Jiang, Ting Zhu, Yu Huang\",\"doi\":\"10.1186/s12985-024-02577-7\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Duck circovirus (DuCV) infections commonly induce immunosuppression and secondary infections in ducks, resulting in significant economic losses in the duck breeding industry. Currently, effective vaccines and treatments for DuCV have been lacking. Therefore, rapid, specific, and sensitive detection methods are crucial for preventing and controlling DuCV.</p><p><strong>Methods: </strong>A lateral flow strip (LFS) detection method was developed using recombinase polymerase amplification (RPA) and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 12a (Cas12a). The RPA-CRISPR/Cas12a-LFS targeted the DuCV replication protein (Rep) and was operated at 37 ℃ and allowed for visual interpretation without requiring sophisticated equipment.</p><p><strong>Results: </strong>The results revealed that the reaction time of RPA-CRISPR/Cas12a-LFS is only 45 min. This method achieved a low detection limit of 2.6 gene copies. Importantly, this method demonstrated high specificity and no cross-reactivity with six other avian viruses. In a study involving 97 waterfowl samples, the Rep RPA-CRISPR/Cas12a-LFS showed 100% consistency and agreement with real-time quantitative polymerase chain reaction.</p><p><strong>Conclusion: </strong>These findings underscored the potential of this user-friendly, rapid, sensitive, and accurate detection method for on-site DuCV detection.</p>\",\"PeriodicalId\":23616,\"journal\":{\"name\":\"Virology Journal\",\"volume\":\"21 1\",\"pages\":\"322\"},\"PeriodicalIF\":4.0000,\"publicationDate\":\"2024-12-19\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11661005/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Virology Journal\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1186/s12985-024-02577-7\",\"RegionNum\":3,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"VIROLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Virology Journal","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1186/s12985-024-02577-7","RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"VIROLOGY","Score":null,"Total":0}
CRISPR/Cas12a and recombinase polymerase amplification-based rapid on-site nucleic acid detection of duck circovirus.
Background: Duck circovirus (DuCV) infections commonly induce immunosuppression and secondary infections in ducks, resulting in significant economic losses in the duck breeding industry. Currently, effective vaccines and treatments for DuCV have been lacking. Therefore, rapid, specific, and sensitive detection methods are crucial for preventing and controlling DuCV.
Methods: A lateral flow strip (LFS) detection method was developed using recombinase polymerase amplification (RPA) and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 12a (Cas12a). The RPA-CRISPR/Cas12a-LFS targeted the DuCV replication protein (Rep) and was operated at 37 ℃ and allowed for visual interpretation without requiring sophisticated equipment.
Results: The results revealed that the reaction time of RPA-CRISPR/Cas12a-LFS is only 45 min. This method achieved a low detection limit of 2.6 gene copies. Importantly, this method demonstrated high specificity and no cross-reactivity with six other avian viruses. In a study involving 97 waterfowl samples, the Rep RPA-CRISPR/Cas12a-LFS showed 100% consistency and agreement with real-time quantitative polymerase chain reaction.
Conclusion: These findings underscored the potential of this user-friendly, rapid, sensitive, and accurate detection method for on-site DuCV detection.
期刊介绍：
Virology Journal is an open access, peer reviewed journal that considers articles on all aspects of virology, including research on the viruses of animals, plants and microbes. The journal welcomes basic research as well as pre-clinical and clinical studies of novel diagnostic tools, vaccines and anti-viral therapies.
The Editorial policy of Virology Journal is to publish all research which is assessed by peer reviewers to be a coherent and sound addition to the scientific literature, and puts less emphasis on interest levels or perceived impact.
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