The Tandem Expression Of The Human/salmon Chimeric CT Gene In E.coli

Calcitonin (CT), a prominent hormone in the thyroid gland of mammal and in ultimobranchial body of fish, is composed of 32 amino acid residues. It's principle function, reducing serum Ca level by suppressing osteoclastic resorption of bones in mammals. Salmon calcitonin (sCT) is already been used in clinical practice, since it's promising hypocalcemic activity which is reported to as high as 40 times than that of human calcitonin.It was synthesized a new human/salmon chimeric CT gene(applied for the patent) in our lab, focussing on retaining hypocalcemic activity of calcitonin while depressing it's antigenicity in the body. In order to enhance the expression quantity, tandem expression method was used. A passage of flexible linker was designed initially, the dimensional structure forecast of SWIIS-MODEL and 3D-PSSM showed, in the tandem CT that be joint by linker, the activity sites and the antigenicity sites could be exposed adequately. Therefore, by the method of isocaudarner, we constructed tandem expression plasmids pLEX-nCT (n=1,2,3,4,5) contained mulyicopy tandem CT genes, which were joint by a passage of DNA coding linker, and expressed them in E.coli GI724. Finnally, SDS-PAGE was used to test the expression product and it was shown that no obvious expression of tandem CT genes. In addition, the result of Western blotting showed that the tandem CT had the antigenicity of human CT. Our works provided experimental base for farther research and application of CT genetic engineering.