Expression And Purfication Of Recombinant Sorcin, Preparation Of Polyclonal Antibody And Their Immunohistochemistry Research

We selected the target gene L249 of encoding sorcin protein from the oral gland of Lampetra japonica cDNA library, next gene L249 was cloned into pET23b and expressed in E.coli BL21, Rosetta BL21 and yeast, then sorcin protein was generated and purified. The purified sorcin protein was used to prepare rabbit-anti-lampetra japonica polyclonal antibody. After using ELISA to determine the potency, the polyclonal antibody of rabbit-anti-lampetra japonica was used as a marker for immunohistochemical detection of the localization of overexpression of sorcin in the main immune organs of lampetra japonica. Preliminarily analysed the mechanism of drug resistance.The expression of protein showed that sorcin was only overexpressed in Escherichia coli BL21 and Rosetta-BL21 as inclusion body. In addition, the amount of expression in Rosetta-BL21 was much than in BL21. Furthermore, sorcin was not expressed in yeast.The concentration determined by BCA was 1.16mg/mL. The potency determined by ELISA was 1:12000.Western-blot analysis of lampetra japonica tissues with sorcin revealed that there is close correlation between the characteristic of histology structure of digestive system and particular feeding behavior. Meanwhile, sorcin was present in a wide variety of tissues, including intestine, heart, liver, germen, oral gland, especially the abundance of the cardiac myocytes. This was relative with sorcin that it could modulate excitation-contraction coupling in the heart. In normal myocardium, sorcin extensively co-localized with ryanodine receptors, whereas in myopathic hearts, the degree of co-localization was markedly disrupted. Several studies have shown that sorcin is association with the cardiac ryanodine receptor (RyR) during the EC of myocardial cells. Our results suggested that sorcin activates Ca2+-ATPase-mediated Ca2+ uptake and restores SR Ca2+ content, and may play critical roles in compensatory mechanisms in both Ca2+ homeostasis and cardiac dysfunction in failing hearts.The results are significant to further understanding much sorcin-involved biologically essential activity and will pave the way for studying the function of sorcin and associated protein.