Construction of antisense RNA expression plasmid for u-PAR and its transfection to highly invasive PC-3M cell subclones.
- Author:
Guoning LIAO
1
;
Qingfen LI
;
Youmei FENG
;
Yaozu DENG
;
Zhuoya LI
;
Feili GONG
;
Ding MA
Author Information
1. Department of Biochemistry and Molecular Biology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030.
- Publication Type:Journal Article
- MeSH:
Cell Line, Tumor;
Cloning, Molecular;
Humans;
Male;
Neoplasm Invasiveness;
Plasmids;
Prostatic Neoplasms;
metabolism;
pathology;
RNA, Antisense;
biosynthesis;
genetics;
Receptors, Cell Surface;
biosynthesis;
genetics;
Receptors, Urokinase Plasminogen Activator;
Reverse Transcriptase Polymerase Chain Reaction;
Transfection;
Urokinase-Type Plasminogen Activator;
antagonists & inhibitors;
genetics;
metabolism
- From:
Journal of Huazhong University of Science and Technology (Medical Sciences)
2003;23(4):369-372
- CountryChina
- Language:English
-
Abstract:
To evaluate the specific inhibition of antisense u-PAR on the u-PAR expressions in highly invasive cell subclones and to determine its blocking function in the invasion by those cells, a cDNA fragment of u-PAR obtained by RT-PCR was inserted into a plasmid vector named pcDNA3 in antisense orientation. Then the antisense u-PAR recombinant was transfected into highly invasive cell subclones. The u-PAR expression in neo-resistant cells was examined by RT-PCR and immunohistochemical assay. Compared to the control cells, the content of mRNA and protein of u-PAR in transfected cells decreased sharply, and the rate of inhibition was 53% and 73%, respectively, indicating that an antisense u-PAR might have played a specific inhibitory role in its expression in the cells, which may provide a good cell model for making further investigation of the inhibitory effects of the antisense u-PAR on invasion in highly invasive cell subclones of human prostate carcinoma.
