人野生型和A53T突变型a-突触核蛋白在SH-SY5Y细胞中表达的相关研究

Objective：To construct the Wild-Type (WT) or A53T mutant α-Synuclein lentiviral expression vector（pEGFP-SNCA-WT and pEGFP-SNCA-A53T）, and transfect them into human neuroblastoma cell neurons (SH-SY5Y),Detect the expressionofα-Synuclein in SH-SY5Y cells,respectively. Methods: The Wild-Type or A53T Mutant α-Synuclein lentiviral expression plasmid was constructed, the orientation of which was confirmed by restriction analysis and was transfected into SHSY5Y cells, respectively. The individual stably transfected colony was subsequently selected in the presence of puromycin, and the gene from transfected SHSY5Y cells was PCR amplified and confirmed by restriction analysis and DNA sequencing. Results: Restriction endonuclease results showed that pEGFP-SNCA-WT and pEGFP-SNCA-A53T plasmids were constructed successfully. The results from antibiotic selection and DNA sequencing showed that the lentiviral vector can be integrated into the genome of SH-SY5Y cell successfully. Western blot assay showed that a-Synuclein could be overexpressed in the transfected SHSY5Y cells successfully. Conclusion: Wild-Type or A53T mutant a-Synuclein lentiviral expression vector was constructed successfully and SH-SY5Y can express a-Synuclein as host cells.The study can lay the foundation for the establishment of Parkinson's disease (PD) model in vitro and the research on the pathogenesis of PD.