Cloning,identification and expression pattern of IFNc in Acipenser sinensis
Chinese sturgeon Acipenser sinensis interferon IFNc(AsFINc)was screened from the transcriptome database,PCR was used for cloning verification,bioinformatics was used to analyze its sequence characteristics and evolutionary status,and qPCR was used to study its expression patterns in different tissues and under different immune stimulation.Results showed that the IFNe cDNA was 549 bp in length and consisted of 183 amino acids.Bioinformatics analysis showed that A.sinensis IFNc belonged to type Ⅰ interferon c subgroup.Phylogenetic tree and homology analysis showed that theIFNe of A.sinensis and Lepisosteus oculatus clustered into one branch.Gene structure analysis showed that the IFNc gene was composed of five exons and four introns,and the intron phase was zero.These differences in protein structure and physicochemical properties suggested that A.sinensis IFNc played different functions in the immune system.IFNc could be expressed in a number of tissues,including liver,blood,cheek,heart,spleen,brain,intestine,muscle,cephalic kidney,mesonephric and skin.The expression level of AsIFNc was the highest in muscle,blood and skin,indicating that AsIFNc might play an important role in muscle,blood and skin.To further understand the function of A.sinensis IFNc,kidney cells were incubated.The experimental groups were added with A.sinensis IFN-γrecombinant protein and Poly I:C,respectively,and the control group was added with the same amount of PBS.After incubation at 28 ℃ for 1 h,8 h,24 h,the medium solution in each hole was removed,and 1 ml Trizol was added to stand at room temperature for 5 min.The solution was collected,total RNA was extracted and converted into cDNA,and then quantitative qPCR detection was performed.The experimental result showed AsIFNc could be induced by poly I:C,and was significantly upregulated with Mx and Viperin,but the upregulation ratio was not as high as Mx and Viperin,and the maximum upregulation ratio was 2.58 times.It was speculated that AsIFNc had certain antiviral function,but its specific antiviral mechanism need further verification.After induction of IFN-γ recombinant protein,AsIFNc,Mx and Viperin were significantly up-regulated between 1-24 h,and the expression levels were the highest at 24 h,with the expression multiple greater than 20.AsIFNc,along with antiviral genes Mx and Viperin,could be induced by IFN-γ recombinant protein in large quantities.A.sinensis IFN-γ could induce the expression of type Ⅰ interferon and cooperate with it to resist foreign pathogens.The expression of A.sinensis IFNc could be up-regulated by viral analogue poly I:C and recombinant A.sinensis IFN γ,indicating that it was involved in antiviral immune regulation as a member of interferon signaling pathway.Herein,AsIFNc was identified as a member of the subgroup interferon IFN-I,with different expression patterns in different tissues.AsIFNc may have certain antiviral ability and resistance to foreign pathogens,but its antiviral ability is weaker than Mx and Viperin and its duration is shorter.IFN-γ can be significantly induced by A.sinensis,and IFN-γ may cooperate with AsIFNc to resist foreign pathogens.Its specific biological function needs to be verified by further experiments.
万方数据
维普
