Localization,Expression And Functional Verification Of Heat Shock Protein 60 In Different Tissues Of Sheep

China is the country with the most abundant sheep breeds in the world.With the continuous growth of the national population and the advancement of production science,there is an increasing demand for sheep meat products.However,the increasingly serious diseases and deteriorating natural conditions have caused major economic losses to,sheep breeding industry.The heat shock protein family(Hsps)is an important anti-infective and anti-stress gene in the organism.HSP60,one of the most conservative large families in Hsps,is widely present in eukaryotes and prokaryotes and is actively involved in the body's immune response.This gene plays an important role in sheeps HSP60 study contributing to the healthy development of China's sheep farming.Using sheep HSP60 gene as a target,Western Blotting and real-time quantitative PCR were used to detect HSP60 protein and gene expression in different tissues of sheeps;the immunohistochemical HSP60 protein localization in different tissues to Fetal cardiomyocytes from 2 to 3-month-old sheeps was used to construct HSP60 overexpression vector pIRES2-EGFP-HSP60.HSP60 was synthesized and silenced to identify the function of gene.The results are as follows:The results showed that:(1)Through the study of gene and protein level,HSP60 was found to be differentially expressed in various tissues and organs of sheeps;HSP60 gene and protein levels in the heart were significantly higher than those in other tissue.It is speculated that HSP60 plays an important role in maintaining the normal function of cardiomyocytes.(2)The HSP60 overexpression vector(pIRES2-EGFP-HSP60)was successfully constructed.The expression of HSP60 gene and protein in overexpression group(pIRES2-EGFP-HSP60)was higher than that in the control group.RT-qPCR detected cell proliferation markers cyclin D1(Cyclin D1),proliferating cell nuclear antigen(PCNA)and anti-apoptotic gene Bcl-2 expression.at 24 h and 48 h.The expression of overexpression group(pIRES2-EGFP-HSP60)was higher than that of the normal group;the expression of proapoptotic gene Bax at 24 h and 48 h overexpression group(pIRES2-EGFP-HSP60)was lower than that in the normal group.(3)Synthetic targeting siRNA silencing HSP60 verified that the level of HSP60 gene and protein expression in the silencing group(SiRNA-HSP60)were lower than those in the control group.The results of RT-qPCR showed that the expression of Cyclin D1,PCNA and Bcl-2 at 24 h and 48 h.The expression of silencing groups(SiRNA-HSP60)was lower than that in the normal group,and the expression of proapoptotic gene Bax in the 24 h and 48 h Silent group(pIRES2-EGFP-HSP60)was higher than that in the normal group.