Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virus
Abstract Background As no treatment or effective vaccine for African swine fever virus (ASFV) is currently available, a rapid, highly sensitive diagnostic is urgently needed to curb the spread of ASFV. Results Here we designed a novel CRISPR-Cas12a based assay for ASFV detection. To detect different...
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| Format: | Article |
| Language: | English |
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BMC
2020-09-01
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| Series: | BMC Microbiology |
| Subjects: | |
| Online Access: | http://link.springer.com/article/10.1186/s12866-020-01966-6 |
| _version_ | 1818351052475334656 |
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| author | Xiaoying Wang Sheng He Na Zhao Xiaohong Liu Yongchang Cao Guihong Zhang Gang Wang Chunhe Guo |
| author_facet | Xiaoying Wang Sheng He Na Zhao Xiaohong Liu Yongchang Cao Guihong Zhang Gang Wang Chunhe Guo |
| author_sort | Xiaoying Wang |
| collection | DOAJ |
| description | Abstract Background As no treatment or effective vaccine for African swine fever virus (ASFV) is currently available, a rapid, highly sensitive diagnostic is urgently needed to curb the spread of ASFV. Results Here we designed a novel CRISPR-Cas12a based assay for ASFV detection. To detect different ASFV genotypes, 19 crRNAs were designed to target the conserved p72 gene in ASFV, and several crRNAs with high activity were identified that could be used as alternatives in the event of new ASFV variants. The results showed that the sensitivity of the CRISPR-Cas12a based assay is about ten times higher than either the commercial quantitative PCR (qPCR) kit or the OIE-recommended qPCR. CRISPR-Cas12a based assay could also detect ASFV specifically without cross-reactivity with other important viruses in pigs and various virus genotypes. We also found that longer incubation times increased the detection limits, which could be applied to improve assay outcomes in the detection of weakly positive samples and new ASFV variants. In addition, both the CRISPR-Cas12a based assay and commercial qPCR showed very good consistency. Conclusions In summary, the CRISPR-Cas12a based assay offers a feasible approach and a new diagnostic technique for the diagnosis of ASFV, particularly in resource-poor settings. |
| first_indexed | 2024-12-13T18:31:36Z |
| format | Article |
| id | doaj.art-567cc7721dae498dba19037122186646 |
| institution | Directory Open Access Journal |
| issn | 1471-2180 |
| language | English |
| last_indexed | 2024-12-13T18:31:36Z |
| publishDate | 2020-09-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Microbiology |
| spelling | doaj.art-567cc7721dae498dba190371221866462022-12-21T23:35:29ZengBMCBMC Microbiology1471-21802020-09-0120111010.1186/s12866-020-01966-6Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virusXiaoying Wang0Sheng He1Na Zhao2Xiaohong Liu3Yongchang Cao4Guihong Zhang5Gang Wang6Chunhe Guo7State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, North Third Road, Guangzhou Higher Education Mega CenterState Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, North Third Road, Guangzhou Higher Education Mega CenterPrecise Genome Engineering Center, School of Life Sciences, Guangzhou UniversityState Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, North Third Road, Guangzhou Higher Education Mega CenterState Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, North Third Road, Guangzhou Higher Education Mega CenterCollege of Veterinary Medicine, South China Agricultural UniversityPrecise Genome Engineering Center, School of Life Sciences, Guangzhou UniversityState Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, North Third Road, Guangzhou Higher Education Mega CenterAbstract Background As no treatment or effective vaccine for African swine fever virus (ASFV) is currently available, a rapid, highly sensitive diagnostic is urgently needed to curb the spread of ASFV. Results Here we designed a novel CRISPR-Cas12a based assay for ASFV detection. To detect different ASFV genotypes, 19 crRNAs were designed to target the conserved p72 gene in ASFV, and several crRNAs with high activity were identified that could be used as alternatives in the event of new ASFV variants. The results showed that the sensitivity of the CRISPR-Cas12a based assay is about ten times higher than either the commercial quantitative PCR (qPCR) kit or the OIE-recommended qPCR. CRISPR-Cas12a based assay could also detect ASFV specifically without cross-reactivity with other important viruses in pigs and various virus genotypes. We also found that longer incubation times increased the detection limits, which could be applied to improve assay outcomes in the detection of weakly positive samples and new ASFV variants. In addition, both the CRISPR-Cas12a based assay and commercial qPCR showed very good consistency. Conclusions In summary, the CRISPR-Cas12a based assay offers a feasible approach and a new diagnostic technique for the diagnosis of ASFV, particularly in resource-poor settings.http://link.springer.com/article/10.1186/s12866-020-01966-6African swine fever; CRISPR-Cas12a; nucleic acid detection; diagnosis; qPCR |
| spellingShingle | Xiaoying Wang Sheng He Na Zhao Xiaohong Liu Yongchang Cao Guihong Zhang Gang Wang Chunhe Guo Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virus BMC Microbiology African swine fever; CRISPR-Cas12a; nucleic acid detection; diagnosis; qPCR |
| title | Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virus |
| title_full | Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virus |
| title_fullStr | Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virus |
| title_full_unstemmed | Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virus |
| title_short | Development and clinical application of a novel CRISPR-Cas12a based assay for the detection of African swine fever virus |
| title_sort | development and clinical application of a novel crispr cas12a based assay for the detection of african swine fever virus |
| topic | African swine fever; CRISPR-Cas12a; nucleic acid detection; diagnosis; qPCR |
| url | http://link.springer.com/article/10.1186/s12866-020-01966-6 |
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