Construction of eukaryotic expression vector of short hairpin RNA for transforming growth factor-beta1
Wang, J.; Wu, J.-Z.; Guo, F.-P.; Zhu, X.-L.; Wen, D.-S.
Hua Xi Kou Qiang Yi Xue Za Zhi 24(2): 113-116
2006
ISSN/ISBN: 1000-1182 PMID: 16704085 Document Number: 597661
To construct the plasmid containing short hairpin RNA (shRNA) of TGF-beta1 expression vector. Short chain oligonucleotide was designed according to the TGF-beta1 mRNA sequence provided by Genebank, then DNA segment was gained through annealing after chemosynthesis, and then was cloned to pWH1 vector. The recombinant TGF-beta1 shRNA expression vector was evaluated by using enzyme cutting. At last, the constructed TGF-beta1 expression vector was transfected into salivary gland mucoepidermoid carcinoma (Ms) cells by Lipofectomine TM 2000, and its effect on TGF-beta1 expression was observed by RT-PCR and immunohistochemistry. Successful construction was identified by enzyme cutting and the constructed plasmid was called pWH1-TGF-beta1. The shRNA and it inhibited the TGF-beta1 mRNA and protein expression effectively. The constructed TGF-beta1 shRNA expression vector can block the TGF-beta1 expression in salivary gland mucoepidermoid carcinoma cells.