Construction of small interfering RNA targeting heparanase gene and its inhibitory effect on the in-vasiveness of human malignant melanoma cell line A375 in vitro
- VernacularTitle:乙酰肝素酶-siRNA表达载体的构建及其抑制黑素瘤细胞株体外侵袭能力的观察
- Author:
Xiao-Yan LIU
;
Hong FANG
;
Zheng-Gang YANG
;
Li-Ming RUAN
;
De-Ren FANG
;
Ying-Guo DING
;
Yi-Na WANG
;
Yu ZHANG
;
Xiao-ling JIANG
;
- Publication Type:Journal Article
- Keywords:
RNA interference;
Gene express;
Melanoma;
Heparanase
- From:
Chinese Journal of Dermatology
2003;0(11):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct the small interfering RNA (siRNA) targeting heparanase gene and its expressing vector,and to observe its interference effect on the expression of heparanase gene and inhibitory effect on the invasive potential of human malignant melanoma A375 cells.Methods Three siRNAs were designed.The recombinant plasmid pRNATU6.1/heparanase-siRNA was designed and constructed. A375 cells were cultured,and transfected with pRNATU6.1/heparanase-siRNA.The cells treated with lipo- fectamine or Opti-MEM served as the controls.Real-time fluorescence quantitative PCR and Western blot were performed to evaluate the expression of heparanase RNA and protein in these treated A375 cells.The in vitro invasive potential of treated A375 cells was assessed by Matrigel gel assay.Results The siRNA targeting heparanase gene was successfully cloned to the eukaryotic expressing vector pRNATU6.1.The expression levels of both heparanase RNA and protein decreased significantly in siRNA-transfected A375 cells than those in the control cells.The in vitro invasive potential of siRNA-transfected cells was also signifi- cantly inhibited as compared with that of the control cells (P
