1.College of Fisheries and Life Science, Dalian Ocean University, Dalian 116023,China;2.Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province,Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou 510380,China
Abstract: In this paper, heat-shock protein(HSP70)gene ORF (open reading frame),as an important medium molecule for change in environmental temperature, was cloned in parasiteIchthyophthiriusmultifiliis, and the expression of the gene was studied in theront,trophont and tomont of the parasite at water temperature of 20, 25, and 30 ℃ by RT-PCR. It was found that Im HSP70 gene ORF was 1995 bp encoding a putative protein of 664 amino acid residues. The gene structure analysis revealed that the Im HSP70 was primarily comprised of α-helix and randon coil-based in secondary structure, and the N-terminal ATPase domain and C-terminal peptide-binding domain in spatial configuration. Comparison with other known HSP70 protein indicated that the Im HSP70 had the maximal similarity (78.08% identity) to that inCarchesiumpolypinum. Phylogenetic analysis revealed thatI.multifiliiswasclustered into a branch withC.polypinum,TetrahymenathermophilaandSterkiellahistriomuscorumby the Im HSP70 gene. The fluorescence quantitative RT-PCR showed that the minimal expression levels of Im HSP70 were observed in theront stage at all the experimental temperature, significantly higher level in trophont than that in theront and tomont at 20 ℃ and 25 ℃(P<0.05),and the maximal level in tomont at 30 ℃. There were higher expression levels in theront and trophont stages in 30 ℃ than that in 25 ℃(P<0.05). Even at 30 ℃,theI.multifiliisstrain showed relatively higher gene expression level and had infectivity to the fish. We can speculate that HSP70 may play an important role in fighting against the external environment temperature change due to the parasite belonging to the tropical type.
2014, Vol. 29 
