Construction and detection of expression vectors of microRNA-9a in BmN cells

文献类型： 外文期刊

第一作者： Huang, Yong

作者： Huang, Yong;Wang, Sheng-peng;Tang, Shun-ming;Zhang, Guo-zheng;Shen, Xing-jia;Zou, Quan;Huang, Yong;Wang, Sheng-peng;Tang, Shun-ming;Zhang, Guo-zheng;Shen, Xing-jia;Huang, Yong

作者机构：

关键词： miRNA-9a (miR-9a);EGFP gene;Bombyx mori N (BmN) Cells;Expression vector

期刊名称：JOURNAL OF ZHEJIANG UNIVERSITY-SCIENCE B （ 影响因子：3.066； 五年影响因子：3.057 ）

ISSN： 1673-1581

年卷期： 2011 年 12 卷 7 期

页码：

收录情况： SCI

摘要： MicroRNAs (miRNAs) are small endogenous RNAs molecules, approximately 21-23 nucleotides in length, which regulate gene expression by base-pairing with 3' untranslated regions (UTRs) of target mRNAs. However, the functions of only a few miRNAs in organisms are known. Recently, the expression vector of artificial miRNA has become a promising tool for gene function studies. Here, a method for easy and rapid construction of eukaryotic miRNA expression vector was described. The cytoplasmic actin 3 (A3) promoter and flanked sequences of miRNA-9a (miR-9a) precursor were amplified from genomic DNA of the silkworm (Bombyx mori) and was inserted into pCDNA3.0 vector to construct a recombinant plasmid. The enhanced green fluorescent protein (EGFP) gene was used as reporter gene. The Bombyx mori N (BmN) cells were transfected with recombinant miR-9a expression plasmid and were harvested 48 h post transfection. Total RNAs of BmN cells transfected with recombinant vectors were extracted and the expression of miR-9a was evaluated by reverse transcriptase polymerase chain reaction (RT-PCR) and Northern blot. Tests showed that the recombinant miR-9a vector was successfully constructed and the expression of miR-9a with EGFP was detected.

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