Cloning and expression analysis of cysteinyl aspartate specific proteinase 3 gene(caspase-3)in Channa argus

【Objective】Cloning and expression analysis of cysteinyl aspartate specific proteinase-3 gene(caspase-3) of Channa argus were conducted,which provided a theoretical basis for exploring the role of this gene in the immune response of C. argus and its role in the immunity of other fishes.【Method】According to the genome information of C. arguspublished by NCBI,the caspase-3 gene sequence was cloned and the deduced caspase-3 amino acid sequence bioinformatics analysis was performed by real-time fluorescence quantitative PCR to detect the expression distribution of caspase-3 gene in various tissues of healthy C. argus and the temporal expression under different stimuli(Seriola quinqueradiata Nocardia seriolae,lipopolysaccharide(LPS) and polymyxcytidylic acid(PolyI:C). 【Result】The caspase-3 gene sequence was 855 bp and encoded 284 amino acid residues,with a protein molecular mass of 31.069 kD and a theoretical isoelectric point (pI) of 6.17. The caspase-3 protein contained 1 CASc domain. Multiple sequence alignment showed that the amino acid sequence of caspase-3 of C. argus and thecaspase-3 of Lutjanus peru,reaching 81.7%. Phylogenetic analysis showed that caspase-3 of C. argus and caspase-3 of L. peru clustered into a single lineage. The results of real-time fluorescence quantitative PCR showed that,the caspase-3 gene of C. argus in the tested tissue(muscle,skin,gills,heart, thymus,brain,head kidney,spleen,liver and blood) were expressed. The spleen and head kidney had the highest expression. Upon stimulation with the different stimuli,the expression level of caspase-3 gene in the spleen,head kidney, blood and liver tissues of C. argus all changed significantly(P<0.05);After incubation of head kidney leukocyte with S. quinqueradiata N. seriolae,LPS and PolyI:C,the caspase-3 gene expression was all significantly increased,and it had an obvious time-dependence.【Conclusion】The caspase-3 gene of C. argus may play an important role in the immune response against bacterial and viral invasion.