Eukaryotic expression of Bm86 gene of Boophilus microplus in Pichia pastoris

A Bm86 gene was cloned from larvae of Boophilus microplus, then inserted into a secretory expression vector pPIC9K, and the expression vector pPIC9K-Bm86 was constructed successfully. The recombinant was linearized with SacⅠ and electroporated into the yeast cell GS115. The multi-copy recombinant P.pastoris strains were screened by G418 and induced by methanol. The expressed product was analyzed by SDS-PAGE and Western-blotting tests. Results showed that Bm86 gene was expressed successfully at 48th hour.The concentration of expressed Bm86 protein was 0.36mg/mL and its molecular weight is approximately 68ku.The expressed Bm86 protein reached up to 32% of total proteins in culture supernatant and could be specifically recognized by the positive serum of rabbit infected by B.microplus.