LAMP assay coupled with CRISPR/Cas12a system for portable detection of
African swine fever virus
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.