Abstract: A thioredoxin reductase (trxR) gene was cloned by PCR amplification from the chromosomal DNA of bacteriumVibrioharveyiSF1. The ORF of the TrxR gene was shown to consist of 960 base pairs, encoding a polypeptide of 319 amino acids. Sequence analysis showed that the amino acid sequence was found 99% similarity with thioredoxin reductases ofVibrioharveyi， 98% withVibrioalginolyticus， 96% withVibrioparahaemolyticus，94% withVibriosplendidusand 92% withVibriofluvialis. A recombinant plasmid pET28a(+)/TrxR was constructed by inserting thetrxR gene into pET28a(+). The recombinant plasmid was further transferred intoEnterobactercoliBL21(DE3) and expressed with IPTG induction. The protein was expressed and purified by Ni(+)-affinity chromatography, and showed a molecular weight of 34 000 on SDS-PAGE. Turbot was immunized with 50 μg of the recombinant protein and the immunized fish was challenged with 0.1 mL ofV.harveyi(3.9×108CFU/mL) for four weeks. The protein vaccine were assessed by ELISA at different days post-vaccination. The antibody titer was found to be 1∶128. The fish in the immunized group had relative percentage survivals (RPS) of 75%. Significant specific antibody responses were detected in the vaccinated fish by indirect ELISA.
2012, Vol. 27 
