Construction of survivin eukaryotic expression vector and its expression in SHG44 cells
AIM: To clone coding sequence of human survivin (SVV) gene, to construct its eukaryotic expression vector and to observe its expression in human brain astrocytoma cell line SHC44. METHODS: By RT-PCR, the specific coding sequence fragments of the SW cDNA were amplified and cloned into eukaryotic expression vector pcDNA3.1. The eukaryotic expression vector pcDNA3.1-SVV was constructed and the gene transfection mediated by the lipofectin was used to introduce pcDNA3.1-SW into human brain astrocytoma cell line SHG44. The stable transfectant cells were established through selecting with G418. Expression of mRNA and protein of survivin was examined by RT-PCR, Westem blot and immunocytochemistry. RESULTS: The SW gene eukaryotic expression vector pcDNA3.1-SW was constructed successfully, which was identified by restriction endonuclease and DNA sequence analysis. Expression of SW gene in SHG44 was demonstrated by RT-PCR,Western blot and immunohistochemistry. CONCLUSION: Survivin gene has been successfully cloned and expressed in SHG44, which lays the experimental foundation for further study of biological functions and mechanisms of SW in SHG44 cells.
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维普
