The wild-type alpha-synuclein over-expression to induce the protein aberrant aggregation of alpha-synuclein in HEK293 cells in vitro.
- Author:
Tao CHEN
1
;
Bei-sha TANG
;
Xiao-ping LIAO
;
Xin-xiang YAN
;
Ru-xu ZHANG
;
Yu-hu ZHANG
;
Jian-guang TANG
;
Li CAO
;
Ji-feng GUO
;
Jing LI
Author Information
- Publication Type:Journal Article
- MeSH: Cells, Cultured; Gene Expression; Humans; Immunohistochemistry; Inclusion Bodies; metabolism; Lewy Bodies; metabolism; Parkinson Disease; genetics; metabolism; alpha-Synuclein; genetics; metabolism
- From: Chinese Journal of Medical Genetics 2006;23(1):19-22
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo investigate over-expression of wild-type alpha-synuclein inducing the aberrant aggregation of alpha-synuclein in HEK293 cell in vitro.
METHODSThe cDNA encoding the human alpha-synuclein without the stop code was cloned into PGEM T-easy vector. Using enzyme map and DNA sequencing analyzed and determined the recombinant plasmid, and then sub-clone the alpha-synuclein cDNA fragment into pEGFP-N1 vector. The recombinant plasmids alpha-synuclein-pEGFP were transfected into HEK293 cells by lipofectamin 2000. The aberrant aggregation of alpha-synuclein was measured by EGFP fluorescence, anti-alpha-synuclein immunocytochemistry. The inclusions in the cultured cells were identified with HE staining.
RESULTSThe restriction enzyme map suggested that eukaryotic expression vector for human wild-type alpha-synuclein gene was constructed successfully. By EGFP fluorescence, anti-alpha-synuclein immunocytochemistry, it could be observed that the alpha-synuclein protein could aggregate in cytoplasm and the Lewy body-like inclusions found in cytoplasm of cultured cells.
CONCLUSIONThe over-expression of wild-type alpha-synuclein can induce protein aberrant aggregation and Lewy body-like inclusions formation in cytoplasm of HEK293 cell in vitro.
