抑制miR-327表达的重组腺病毒载体的构建及鉴定

Construction and Identification of Recombinant Adenovirus Vector Inhibiting miR-327 Expression

目的:构建并鉴定可稳定抑制miR-327表达的重组腺病毒载体。方法:根据miR-327成熟体反向互补序列,合成寡核苷酸链,构建抑制miR-327表达的质粒载体,并对阳性克隆子进行测序鉴定。miR-327shRNA重组质粒载体经包装、扩增、纯化获得miR-327shRNA重组腺病毒,测定滴度。结果:获得最终滴度为1×10^(11) PFU/mL的大鼠miR-327shRNA重组腺病毒载体。结论:成功构建具备一定滴度的大鼠miR-327shRNA重组腺病毒载体,为后期揭示miR-327在心血管疾病中的作用及机制提供了基础。

Objective:To construct and identify recombinant adenoviral vectors that inhibits the expression of miR-327.Methods:Complementary oligonucleotide chains were synthesized on the basis of mature miR-327 sequences.Plasmid vectors inhibiting miR-327 expression were constructed,and positive clones were sequenced.The recombinant plasmid vectors inhibiting miR-327 expression were packaged,amplified and purified.The titer of obtained recombinant adenoviral vectors that inhibits the expression of miR-327 was further determined.Results:Recombinant adenovirus vectors carrying with miR-327 shRNA were successfully constructed,with the final titer of 1×1011 PFU/mL.Conclusion:The recombinant adenovirus vectors carrying with miR-327 shRNA are successfully constructed,which provides a basis for revealing the role and mechanism of miR-327 in cardiovascular diseases.