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LAMP assay coupled with CRISPR/Cas12a system for portable detection of African swine fever virus
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  • Bo YANG,
  • zhengwang shi,
  • Yuan Ma,
  • Lijuan Wang,
  • Liyan Cao,
  • Juncong Luo,
  • Ying Wang,
  • Rui Song,
  • Yiyong yan,
  • kehu yuan,
  • Hong Tian,
  • Haixue Zheng
Bo YANG
State Key Laboratory of Veterinary Etiological Biology
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zhengwang shi
State Key Laboratory of Veterinary Etiological Biology
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Yuan Ma
State Key Laboratory of Veterinary Etiological Biology
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Lijuan Wang
State Key Laboratory of Veterinary Etiological Biology
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Liyan Cao
State Key Laboratory of Veterinary Etiological Biology
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Juncong Luo
State Key Laboratory of Veterinary Etiological Biology
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Ying Wang
State Key Laboratory of Veterinary Etiological Biology
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Rui Song
State Key Laboratory of Veterinary Etiological Biology
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Yiyong yan
Research and Development Department Shenzhen Bioeasy Biotechnology Co Ltd Shenzhen Guangdong 518101 China
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kehu yuan
Research and Development Department Shenzhen Bioeasy Biotechnology Co Ltd Shenzhen Guangdong 518101 China
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Hong Tian
State Key Laboratory of Veterinary Etiological Biology
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Haixue Zheng
State Key Laboratory of Veterinary Etiological Biology
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Abstract

African swine fever (ASF) is one of the most severe infectious diseases of pigs. In this study, a LAMP assay coupled with the CRISPR Cas12a system was established in one tube for the detection of the ASFV p72 gene. The single-strand DNA-fluorophore-quencher (ssDNA-FQ) reporter and CRISPR-derived RNA (crRNAs) were screened and selected for the CRISPR detection system. In combination with LAMP amplification assay, the detection limit for the LAMP-CRISPR assay can reach 7 copies/μl of p72 gene per reaction. Furthermore, this method displays no cross-reactivity with other porcine DNA or RNA viruses. The performance of the LAMP-CRISPR assay was compared with real-time qPCR tests for clinical samples, a good consistency between the LAMP-CRISPR assay and real-time qPCR was observed. The method shed a light on the convenient, portable, low cost, highly sensitive and specific detection of ASFV, demonstrating a great application potential for monitoring on-site ASFV in the field.

Peer review status:ACCEPTED

07 May 2021Submitted to Transboundary and Emerging Diseases
07 May 2021Submission Checks Completed
07 May 2021Assigned to Editor
10 May 2021Reviewer(s) Assigned
24 Jun 2021Review(s) Completed, Editorial Evaluation Pending
28 Jun 2021Editorial Decision: Revise Minor
01 Jul 20211st Revision Received
01 Jul 2021Submission Checks Completed
01 Jul 2021Assigned to Editor
03 Jul 2021Reviewer(s) Assigned
07 Aug 2021Review(s) Completed, Editorial Evaluation Pending
07 Aug 2021Editorial Decision: Accept