摘要
目的利用双分子荧光互补(BiF C)分析检测α-突触核蛋白的聚集及在细胞间的传播。方法首先利用高灵敏的荧光素酶human Gaussiaprinceps luciferase作为报告蛋白,将其N端(1～93)与C端(94～169)分别与α-突触核蛋白的C端融合,记作L1和L2,并将L1和L2同时在SK-N-SH细胞中共同表达,结果显示BiFC分析可以特异地检测出α-突触核蛋白的聚集。接下来将L1和L2分别在SK-N-SH细胞中表达,收集L1的培养基加入到表达L2的细胞中。同理,收集L2的培养基加入到表达L1的细胞中。结果 BiFC分析可检测到α-突触核蛋白在细胞间传播。结论 BiFC可有效检测α-突触核蛋白的聚集及在细胞间传播。
Objective To detect the aggregation and transmission of the α-synuclein by bimolecular fluorescence complementation (BiFC) assays. Methods Human Gaussiaprinceps luciferase luciferase (hGLuc) was used as a reporter protein. The N (1 ~ 93) or C (94~169) terminal of h GLuc was fused with α-synuclein C terminal,named L1 and L2. Then the two kinds of fusion protein were co-ex-pressed in SK-N-SH cells. Results BiFC assays could detect the aggregation of the α-synuclein. Subsequently,L1 and L2 were expressed inSK-N-SH cells respectively. Then the medium of L1 and L2 were exchanged with each other. BiFC assays could detect the transmission of theα-synuclein.Conclusions BiFC assays could detect the aggregation and transmission of the α-synuclein sensitively and effectively.
作者
任晓曦
赵云
韩玉坤
郑焱
杨慧
张建亮
REN Xiao-Xi;ZHAO Yun;HAN Yu-Kun(Department of Neurobiology,School of Basic Medical Sciences,Capital Medical University,Center for Parkinson's Disease,Beijing Institute for Brain Disorder,Key Laboratory for Neurodegenerative Disease of Ministry of Education,Beijing,100069,China)
出处
《中国老年学杂志》
CAS
北大核心
2018年第23期5769-5772,共4页
Chinese Journal of Gerontology
基金
国家自然科学基金(31571202
31271136)
北京市属高等学校青年拔尖人才培养计划(CIT&TCD201504087)
北京市教委科技计划(KZ201210025020)
关键词
Α-突触核蛋白
聚集
传播
双分子荧光互补分析法
荧光素酶
α-synuclein
Aggregation
Transmission
Bimolecular fluorescence complementation assays
Fluorescent enzyme
