Construction of signal peptide-canstatin expression vector and its secretable expression in Eca-109 cells
- VernacularTitle:信号肽-canstatin真核表达载体的构建及其在Eca-109细胞中的分泌表达
- Author:
Weihong HOU
;
Fang TIAN
;
Jianmin WANG
;
Zhichao WANG
;
Huayan CHEN
;
Lexun XUE
- Publication Type:Journal Article
- Keywords:
Canstatin;
Angiogenesis inhibitors;
Esophageal neoplasms;
Eca-109 cells
- From:
Chinese Journal of Pathophysiology
2000;0(10):-
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To construct signal peptide-canstatin expression vector pEGFP-C1-SP-Can and express secretable mouse canstatin fusion protein in Eca-109 cells.METHODS: Site-directed mutagenesis was used in amplifying the signal peptide of murine plasminogen to construct the plasmid pEGFP-C1-SP.The cDNA of mouse canstatin,obtained from a cloning vector pMD18T-Can by PCR,was inserted into pEGFP-C1-SP to construct a secretable expression vector pEGFP-C1-SP-Can.Constructed plasmid pEGFP-C1-SP-Can was transiently transfected into Eca-109 cells via lipofectamine,and subsequently its secretable expression in the medium of cultured Eca-109 was observed by Western blotting.RESULTS: DNA sequencing and restriction enzyme analysis attested the validity of the constructed plasmids pEGFP-C1-SP and pEGFP-C1-SP-Can.EGFPcanstatin fusion protein was proved to be secretably expressed in Eca-109 by Western blotting.CONCLUSION: It is concluded that the constructed vector pEGFP-C1-SP-Can is valid and capable of expression in Eca-109,these findings provide a basis for testing the function of mouse canstatin and its application in gene therapy.
