Development of a point-of-care testing method for Feline Coronavirus using CRISPR techniques

Its high genetic variability challenges existing molecular diagnostic techniques in clinical environments. Thus, we developed visual assays for the detection of FCoV nucleic acid based on CRISPR-Cas13a trans-cleavage activity.

Recombinant plasmid DNA and CRISPR RNAs (crRNAs) were synthesized, respectively, targeting to FCoV RdRp, S, E and N genes of FCoV. Additionally, the Cas13a protein was purified through an Escherichia coli prokaryotic expression system. To validate the efficacy of the FCoV-CRISPR-Cas13a strip, five distinct reaction systems comprising various components were evaluated, with visual outputs assessed via fluorescence detection and colloidal gold immunochromatographic test strips.