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作 者 ：彭特, 王丙云, 李东升, 陈志胜, 陈胜锋, 计慧琴, 陈金顶

期 刊 ：生物技术通报 2014年 0卷 5期 页码：117-121

关键词：鸡；胚胎干细胞；cNanog；cPouV；干扰载体；

Keywords:Chicken Embryonic, Stem, Cells, CNanog, CPouV Interference vector,

摘 要 ：旨在构建靶向鸡胚胎干细胞多能性相关基因cNanog和cPouV的siRNA表达载体，并在体外检测其对cNanog和cPouV表达的抑制作用。分别选择cNanog和cPouV中3个区域合成寡聚核苷酸构建siRNA表达载体，同时构建cNanog和cPouV全基因表达载体来检测基因表达下调的效果。经Western blotting检测显示，所构建的siRNA表达载体可在293T细胞内表达siRNA或shRNA，诱发RNA干扰效应，从而抑制目的基因cNanog和cPouV的表达。不同位点的寡核苷酸序列抑制效率不同，其中pS3-cNanog和pS3-cPouV重组质粒的干扰效果和抑制效率最好。研究结果为后续得到稳定干扰cNanog 和cPouV基因的重组载体和进一步研究cNanog 和cPouV基因功能奠定了基础。

Abstract:The study was to construct the siRNA(small interfering RNA)expression vectors targeting cNanog and cPouV of Chicken Embryonic Stem cells(CES cells), and evaluate its inhibitory effect on cNanog and cPouV in 293T cells. Three regions of cNanog and cPouV gene were respectively selected to synthesize oligonucleotides, and construct interference vectors. These vectors were respectively transfected into 293T cells as whole-gene expression vectors were transduced simultaneously. Western blotting analysis showed target genes expression were downregulated after RNAi and the oligonucleotide sequence S3 was found to be the most efficient one in interfering effect. The results of this experiment indicate siRNA or shRNA(short hairping RNA)were translated into cells and suppress the expression of target genes.

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