CLONING AND EXPRESSION OF CAPRINE INTERFERON-GAMMA

Caprine interferon-gamma (IFN-gamma) cDNA was cloned from mitogen stim ulated peripheral blood mononuclear cell (PBMC) RNA utilizing the reve rse transcription-polymerase chain reaction (RT-PCR). The cDNA open re ading frame (ORF) is 498 bp, encoding a putative 166 amino acid (aa) p rotein (19 327 Da). The predicted aa sequence homology of caprine IFN- gamma and the corresponding ovine, bovine and cervine cytokine is 98.8 %, 95.2% and 92.8%, respectively. IFN-gamma cDNA was subcloned and exp ressed in two different plasmids under the control of either the human cytomegalovirus (CMV) immediate early promoter or the caprine arthrit is-encephalitis virus long terminal repeat (CAEV LTR). Recombinant cap rine IFN-gamma (rCaIFN-gamma) secreted by transfected COS-7 cells shar ed at least two antigenic epitopes with recombinant bovine IFN-gamma ( rBoIFN-gamma) and exhibited biological activity in the vesicular stoma titis virus (VSV) cytopathic effect reduction assay. In-vivo expressio n of IFN-gamma cDNA promoted by the CAEV LTR was confirmed by the intr amuscular (IM) injection of Balb/C mice with plasmid followed by Weste rn blot analysis of mouse serum against purified rCaIFN-gamma produced in E. coli. (C) 1998 Elsevier Science B.V.