OVEREXPRESSION AND PURIFICATION OF RECOMBINANT NUCLEOPROTEIN (N) OF RABIES VIRUS IN E. COLI

Abstract

In order to obtain the recombinant nucleoprotein to develop rabies diagnostic test and recombinant rabies vaccine, nucleoprotein of rabies virus was expressed in recombinant E. coli. The 1.35 kb of nucleoprotein gene was amplified by PCR technique using specific primers. PCR products were digested with BamHI, SaeT and inserted into pET21 a( +) vector which were treated with the same enzymes to form recombinant pET2Ia(+)N vector. This was transformed into E. coli Rosetta_gami^TM cells. The expression of N protein gene was regulated by T7 promoter using 1 mM IPTG as an inducer, at 28°C, for 5 hours. The recombinant N protein with the molecular weight of 51 kDa was checked by 12.6% SDS-PAGE and purified by using His-tag affinity column. MALDI-TOF and MS/MS analyses of pep tides  frecombinant N protein showed that amino acids ofthcse peptides perfectly matched with those ofN protein in NCBI protein Database.