Prokaryotic expression and identification of nucleocapsid gene of porcine epidemic diarrhoea virus

To clone and express nucleocapsid(N)gene of porcine epidemic diarrhoea virus(PEDV),a pair of primer was designed according to PEDV-N gene which had reported in GenBank.Viral RNA was extracted from the small intestine of piglet affected by PEDV,the N gene was amplified by RT-PCR and cloned into pET-28a-His-Sumo vector.The recombinant vector was confirmed by enzyme digestion and sequencing.The positive plasmid was transformed into competent Escherichia coli BL21,then expressed under IPTG induction.A protein with 60.0ku of molecular weight was confirmed by SDS-PAGE and could specifically react with PEDV positive serum identified by Western-blot.This experiment proved that the N protein of PEDV was successfully expressed in prokaryotic expression system,and the product had good reactinogenicity.