Cloning and prokaryotic expression of goose interleukin-18 gene and the preparation of polyclonal antibody to recombinant goose IL-18

According to duck,turkey and chicken IL-18 gene sequences available in GenBank,a pair of primers was designed and synthesized to clone the middle sequence of goose IL-18(GIL-18) gene about 333bp by RT-PCR from the total RNA of goose spleen lymphocyte.The sequence analysis showed that the nucleotide homology of the middle sequence between goose and duck was the highest 99%.Then according to the duck IL-18 gene sequence,three upstream and downstream primers were designed and a battery testing of RT-PCR was carried out to clone goose full-length IL-18 gene.The results indicated that the goose IL-18 gene sequence about 642bp from three breeds geese in Guangdong were amplified,cloned and sequenced successfully.And the three sequences were the same one,including a ORF about 603bp.The ORF that was 23.2ku in molecular weight encoded 200 amino acids.The leader sequence was 30 amino acids at the N-terminus,and the other 170 amino acids was functional sequence.This sequence was registered in GenBank(EF159728) for the first time.The recombinant expression plasmid pET-GIL-18 was constructed.Then the recombinant with GIL-18 gene was expressed and purified.Based on it,the polyclonal antibody of rabbit anti-rGIL-18 was made.