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《Chinese Journal of Health Laboratory Technology》 2014-08
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The targeted-regulating role of hsa-miR-326 on gene BCL2L1 and BAK1 by dual luciferase reporter system

YU Shi-fang;LI Qiang;HONG Jun-ying;JIN Chen-hua;CHEN Bi-le;The First Affiliated Hospital of Wenzhou Medical University;  
Objective To identify the influence of hsa-miR-326 on the expression of BCL2L1 and BAK1,based on construction of psiCHECK2 luciferase reporter gene vector. Methods The potential fragments of hsa-miR-326 target genes BCL2L1 and BAK1 were predicted by the bioinformatics analyzing tools online. The target fragment of the BCL2L1 and BAK1 and their mutation sequence were connected to psiCHECK-2 vector. The constructs were cotransfected into 293T cells with hsa-miR-326 mimics or negative control( mir-NC) by lipofectamineTM 2000( lipo2000),respectively. Dual luciferase reporter system was used to determine the luciferase activity. Results Luciferase assays revealed that hsa-miR-326 could significantly diminish luciferase activity from the reporter vector containing the 3' UTR of BCL2L1( P 0. 01),While the suppression of luciferase activity was not found in BCL2L1-mut,BAK-1 and BAK1-mut( P 0. 05). Conclusion The results strongly suggest that hsa-miR-326 binds to the 3' UTR of BCL2L1,but not to the 3' UTR of BAK1. hsa-miR-326 might be related to the occurrence and development of storage platelet apoptosis by regulating the expression of the apoptosis related gene BCL2L1.
【CateGory Index】: R346
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