Abstract: According to the data in genebank and the characterization of the codon usage in yeast,the optimized gene encoding SARS nucleocapsid protein(1 296 bp) by mutating low-usage codon into synonymous ones was synthesized.After recombined with the CTL epitope gene(195 bp),the gene was cloned into secreted expression vector pPIC9K to pPIC9K-N.pPIC9K-N was transformed into the Pichia pastoris GS115,and then the positive recombinant GS115/pPIC9K-N was screened by MM and MD plates and identified by PCR further.Nucleocapsid protein was highly expressed with a yield of 20% as a secretion under methanol induction and the expressed nucleocapsid protein was isolated and purified by salting out.SDS-PAGE and Western blotting results showed that both the expressed and purified nucleocapsid protein were recognized specifically by SARS antiserum,showed good reactiongenicity and antigenic specificity.

Key words: Severe acute respiratory syndrome(SARS), Nucleocapsid protein, Pichia pastoris