Expression and localization of adenovirus-mediated transcriptional factor Runx3 in malignant glioblastoma cells / 细胞与分子免疫学杂志

ABSTRACT

AIM: To construct the replicative defecient 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-CMVRunx3 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 Pmel 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. CONCLUSION: 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.