Cloning of a gene fragment encoding chicken complement component C3d with expression and immunogenicity of Newcastle disease virus F gene–C3d fusion protein

Abstract

The gene fragment coding for the complement protein C3 (chC3d) from Arbor Acres (AA) chickens was cloned and expressed as a fusion protein for its application in the vaccine study of chicken, and for in vitro experiments. The chC3d fragment strengthened B-cell responses when complexed with antigen. Three potential vaccine construct units were engineered to contain two, four and six copies of the chC3d-P29 coding gene linked to the F gene of Newcastle disease virus, an economically important pathogen of chicken that is classified as a list A contagious disease of poultry by the Office International des Epizooties. Recombinant chC3d protein and C3d-P29 proteins that contained the F gene of Newcastle disease virus (C3d-F-P29.n) were generated separately in Escherichia coli and chick embryo fibroblast cells with the help of expression vectors. All recombinant proteins were analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and western blotting. Analysis of the immunogenicity of different repeats of C3d-F-P29.n revealed that C3d-P29 had an enhancing effect on the immunogenicity of antigens, and that six or more repeats of C3d-P29 may be necessary for efficient enhancement of antigen-specific immune responses. To date, published research into the adjuvant activities of C3d has been limited to experiments in mice, rabbits and cattle. The adjuvant properties of C3d have not been assessed in poultry using homologous C3d in association with antigens relevant to the target species. The chicken C3d fusion proteins detailed in this study are the first reported and they provide a basis for immunization trials in chicken, studies of receptor binding and cell activation of chicken lymphocytes, and investigations of new types of vaccine, including genetic recombinant and DNA vaccines against various pathogens.