Cloning and Poor Expression of S1 Gene of Infectious Bronchitis Virus (Genotype Variant 2) in pET28a

Avian infectious bronchitis (IB) is an economically important, highly contagious, acute, upper-respiratory-tract disease of chickens and other bird species, caused by the avian coronavirus infectious bronchitis virus (IBV). The aim of this study was the expression of the S1 gene protein genotype variant-2 in pET28a. In vitro protein expression is an important method for obtaining many viral proteins to investigate their protective and biological properties. Since the S1 glycoprotein of the infectious bronchitis virus induces virus-neutralizing antibodies, it is an appropriate candidate for producing a recombinant vaccine against infectious bronchitis disease.

In this study, the S1 gene fragment of infectious bronchitis virus genotype IS/1494/06 was amplified using Reverse Transcriptase-Polymerase Chain Reaction (RT_PCR) and was inserted into the pTG19-T vector. The construct was subcloned into pET28a and inserted into E.coli BL21 competent cells.

The insertion was proved by PCR analysis, and isolation of the S1 gene from the construct was carried out. Finally, it was sequenced. The SDS-PAGE assay showed that a protein about 60 kDa was expressed, but the western blot assay did not confirm the expression of the S1 protein.

S1 protein did not express well. To obtain properly expressed proteins, we suggest trying different expression vectors or choosing smaller important portions of the S1 gene.