[Construction of the eukaryotic expression vector pIRES2-EGFP-Axin and its expression in glioma cells].
OBJECTIVE
To construct the eukaryotic expression vector pIRES2-EGFP-Axin, and to express Axin in C6 glioma cells.


METHODS
The Axin gene was amplified by PCR using pCMV5-HA-Axin as a template, and confirmed by DNA sequencing. The eukaryotic expression vector pIRES2-EGFP-Axin was constructed by introducing Axin DNA fragment into the sites of Nhe I and Sal I of pIRES2-EGFP vector. The plasmid was transfected into the C6 cells using lipofectamine. The expressed EGFP was observed under fluorescent microscope and the Axin protein expression was detected by immunostaining using anti-Axin antibody.


RESULTS
The eukaryotic expression vector pIRES2-EGFP-Axin was constructed and transfected successfully into C6 glioma cells. The green fluorescence of EGFP was observed in the plasma and nuclei of transfected cells, and Axin protein was only found in the plasma.


CONCLUSIONS
The recombinant expression vector pIRES2-EGFP-Axin was constructed, and the EGFP and Axin gene could be co-expressed in the C6 cells. This study laid a foundation for the further research of the function of Axin in cell differentiation, growth and tumorigenesis.
Zhang
Li-ying
LY
Department of Pathology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China. zly79921@sohu.com
Li
Qing
Q
Chen
Guang-sheng
GS
Lin
Sheng-cai
SC
Ye
Jing
J
Si
Shao-yan
SY
Zhang
Feng
F
D000818:Animals; D060466:Axin Protein; D045744:Cell Line, Tumor; D004587:Electrophoresis, Agar Gel; D005057:Eukaryotic Cells / Q000378:metabolism*; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics*; D005910:Glioma / Q000235:genetics / Q000473:pathology*; D049452:Green Fluorescent Proteins / Q000235:genetics*; D051379:Mice; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics*; D014162:Transfection
D060466:Axin Protein; D011994:Recombinant Proteins; D012097:Repressor Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins