1 Department of pharmacognosy, College of pharmacy, Third Military Medical University, Chongqing 400038, China 
2 Department of Oncology, General Hospital of Shenyang Military Region, Shenyang 110016, China 
3 Institute of genome, College of Medicine, Tsinghua University, Beijing 100084, China 
4 Department of Histology and Embryology, College of Medicine, Third Military Medical University, Chongqing 400038, China

Abstract 

Objective 
To construct effective RNA-interference plasmids targeting mouse HIF-1α gene and testify their effects and specificities in interfering HIF-1α expression. 
Methods 
Three RNA-interference plasmids targeting mouse HIF-1α gene, pBS/U6/HIF-1α-siRNAI∼III, were constructed and identified using double digestion method in the present study. RT-PCR, immunostaining and western blotting were employed to detect the expression alterations of HIF-1α in 293T cells following transfections of the three plasmids, respectively. The interference effect of pBS/U6/HIF1αi-II in SH-SY5Y cell line was further investigated. 
Results 
All the three RNA-interference plasmids, especially pBS/U6/HIF1αi-II, showed significant inhibition in HIF-1α expression in 293T cell line. pBS/U6/HIF1αi-II could also inhibit HIF-1α expression in SH-SY5Y cell line, in a dose-dependent way. 
Conclusion 
Plasmid pBS/U6/HIF1αi-II constructed in our study can effectively and specifically inhibit HIF-1α expression, and its role in neural tube development and dysfunction will be further investigated. Construct of pBS/U6/HIF1αi-II plasmid will provide a useful tool to study the role of HIF-1 pathway in embryogenesis, oncogenesis and ischemia development.

Keywords

HIF-1α; RNA interference; pBS/U6/HIF-1α-siRNA

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