The comparison of antisense oligonucleotides、small interferingRNA targeting KDR for gene silencing
- VernacularTitle:KDR为靶的反义寡核苷酸、siRNA活性比较
- Author:
Sujun ZHENG
- Publication Type:Journal Article
- Keywords:
Antisense oligonucleotides;
Small interfering RNA;
KDR
- From:
Journal of Chongqing Medical University
1986;0(03):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate and compare the inhibitory effects on KDR gene expressinon of antisense oligonucleotides(asONs),short hairpin small interfering RNA(siRNA) targeting the mRNA of VEGFR2/KDR(vascular endothelial growth factor receptor 2/kinase insert domain-containing receptor)in MCF-7 cells.Methods:3 antisense oligonucleotides were designed.According to the target KDR mRNA accessible sites of antisense oligonucleotides,3 DNA vector-based pPUR/U6/KDR-siRNA for intracellular transcription KDR specific short hairpin small interfering RNA were constructed.The inhibitory effects on KDR protein expression of antisense oligonucleotides,and pPUR/U6/KDR-siRNA for stable transfection were evaluated using Western-blotting assay.The antisense oligonucleotides inhibitory effects on cell proliferation were determined with MTT assay,while the growth curve of MCF-7 cells stably transfected with pPUR/U6/KDR-siRNA was measured using cell counting.Results:3 antisense oligonucleotides significantly reduced the KDR protein expression,with the inhibitory rates being 66.16%,50.21% and 58.35% at 0.8?mol/L,respectively.They also decreased the proliferation of MCF-7 cells in a dose dependent manner.Compared to the cells transfected with vacant pPUR/U6 control vector,two of three pPUR/U6/KDR-siRNA plasmids caused effcient down-regulation of KDR protein expression in stable transfected MCF-7 cells.At the 15th day after stable transfection of pPUR/U6/KDR-siRNA,the inhibitory rates were 75.21% and 80.08%,respectively;and at the 30th day the inhibitory rates were 59.29% and 60.15%,respectively.The stable transfection of pPUR/U6/KDR-siRNA also slowed MCF-7 cells growth.Conclusion:It is an effective approach to design siRNA according to the antisense oligonucleotides target accessible sites.The endogenous short hairpin siRNA produced by DNA plasmid can more significantly inhibit KDR gene expression than the antisense oligonucleotides for a long time.
