[Expression and localization of adenovirus-mediated transcriptional factor Runx3 in malignant glioblastoma cells].
OBJECTIVE
To construct the replicative deficient adenovirus Ad-Runx3 expressing Runx3, and to express it in U251 malignant glioblastoma cells.


METHODS
The runx3 gene with a flag tag was amplified by PCR using pCMV5-AML2 as a template, and was confirmed by DNA sequencing. The adenovirus shuttle vector pShuttle-CMV-Runx3 was constructed by introducing runx3 DNA fragment into the sites of Kpn I and Xho I of pShuttle-CMV vector. This recombinant plasmid was linearized by PmeI and electronically transfected into BJ5183 cells to get the recombinant adenovirus vector Ad-Runx3. The recombinant adenovirus expressing Runx3 was infected into U251 malignant glioblastoma cells. The expression of exogenous Runx3 was observed by immonoblotting and its localization was detected by immunostaining using anti-Flag tag antibody.


RESULTS
The recombinant adenovirus expressing Runx3 with a Flag tag was constructed and infected into U251 glioblastoma cells. The exogenous Runx3 protein was detected only in the nuclei.


CONCLUSIONS
The recombinant adenovirus expressing Runx3 with a Flag tag is constructed successfully, and the Runx3 protein expressed in the nuclei of infected cells. The study laid a foundation for further research of the function of Runx3 in gliocarcinogenesis.
Bai
Wen-Dong
WD
The State Key Laboratory of Cancer Biology, Xi'an 710032, China.
Zhang
Rui
R
Meng
Yan-Ling
YL
Cao
Yun-Xin
YX
Wang
Tao
T
Zhao
Jing
J
Yang
An-Gang
AG
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D050677:Core Binding Factor Alpha 3 Subunit / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D015870:Gene Expression; D005909:Glioblastoma / Q000473:pathology*; D006801:Humans; D051379:Mice; D010957:Plasmids / Q000235:genetics; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
D050677:Core Binding Factor Alpha 3 Subunit; D011993:Recombinant Fusion Proteins