抑制CDC25A表达的siRNA真核表达载体的构建

Construction of eukaryotic expression vector of siRNA-inhibited CDC25A expression

目的在哺乳动物细胞中构建并鉴定抑制CDC25A表达的siRNA真核表达载体。方法根据CDC25A的基因序列,设计特异性的siRNA,将合成的siRNA核酸片段退火形成双链后连接到经Bam HI和HindⅢ双酶切后的psilencer4.1真核表达载体,命名为psilencer4.1-CDC25A以及psilencer4.1-Control,并进行酶切及测序鉴定。结果通过酶切鉴定和序列测定,成功地构建出抑制CDC25A表达的siRNA载体及其阴性对照载体。结论成功构建和验证了siRNA真核表达载体,为下一步研究奠定了良好的基础。

Objective To construct and identify an eukaryotic expression vector of siRNA-inhibited CDC25A expression in mammalian cells. Methods CDC25A-specific siRNA was designed and synthesized according to the sequence of CDC25A. After annealing the nucleic acid fragment of the siRNA to form the double-stranded, they were connected with the eukaryotic expression vector psilencer4. 1 which was pre-digested with Barn HI and Hind Ⅲ. Then the constructed vectors, named psilencer4. 1-CDC25A and psilencer4. 1-Control, were digested and sequenced. Results The vector of siRNA-inhibited CDC2.SA expression and the negative control vector were constructed, sequenced and identified. Conclusion The successful construction and validation of the siRNA-inhibited CDC25A expression vector lays a good foundation for the next steps in CDC25A-related study.