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张玫, 何訸, 宋昊岚等. 双荧光素酶报告基因系统鉴定has-miR-577和has-miR-583对FGF-21基因的靶向调控[J]. koko体育app 学报(医学版), 2016, 47(6): 916-919.
引用本文: 张玫, 何訸, 宋昊岚等. 双荧光素酶报表基因组组程序鉴定会has-miR-577和has-miR-583对FGF-21基因组组的靶向疗法自我调节[J]. 上海本科大学学报(医疗版), 2016, 47(6): 916-919.
ZHANG Mei, HE He, SONG Hao-lan. et al. The Targeted Regulating Role of Has-miR-577 and Has-miR-583 on Gene FGF-21 Detected by the Dual[J]. Journal of Sichuan University (Medical Sciences), 2016, 47(6): 916-919.
Citation: ZHANG Mei, HE He, SONG Hao-lan. et al. The Targeted Regulating Role of Has-miR-577 and Has-miR-583 on Gene FGF-21 Detected by the Dual[J]. Journal of Sichuan University (Medical Sciences), 2016, 47(6): 916-919.

双荧光素酶报告基因系统鉴定has-miR-577和has-miR-583对FGF-21基因的靶向调控

The Targeted Regulating Role of Has-miR-577 and Has-miR-583 on Gene FGF-21 Detected by the Dual

  • 摘要: 目的 通过构建成纤维细胞生长因子-21基因(fibroblast growth factor 21,FGF-21)荧光素酶报告基因载体,观察has-miR-577和has-miR-583对FGF-21基因的靶向调控。方法 利用生物学信息网站对has-miR-577和has-miR-583靶向结合FGF-21 3′UTR区的位点进行分析,设计合成包含与has-miR-577和has-miR-583结合序列及其突变序列的FGF-21基因片段,构建野生型(psiCHECK2-FGF-21)和突变型(psiCHECK2-FGF-21-mut)FGF-21双荧光素酶报告基因载体。进行双酶切电泳和测序鉴定后,FGF-21野生型与突变型双荧光素酶报告基因载体分别+〔hsa-miR-577模拟物、hsa-miR-583模拟物和miR无义序列阴性对照 (miR negative control,miR-NC)〕转染293T 细胞,检测荧光素酶活性,观察has-miR-577和has-miR-583对FGF-21表达的影响。结果 双酶切电泳和测序结果显示,野生型(psiCHECK2-FGF-21)和突变型(psiCHECK2-FGF-21-mut)基因载体片段大小、序列结果与实验预期一致,载体构建成功。has-miR-577和has-miR-583可抑制野生型FGF-21载体的荧光表达(PFGF-21载体的荧光表达无抑制作用。结论 has-miR-577和has-miR-583可以靶向调控FGF-21的表达。  
    Abstract: Objective To determine the targeted regulating role of has-miR-577 and has-miR-583 on the expression of fibroblast growth factor 21 (FGF-21) based on a constructed luciferase reporter FGF-21 gene vector. Methods The site of has-miR-577 and has-miR-583 target genes FGF-21 were predicted by the bioinformatics analyzing tools online.FGF-21 gene fragments, combined with has-miR-577 or has-miR-583 sequences and mutant sequences,were designed and synthesized.The wild type (psiCHECK2-FGF-21) and mutant (psiCHECK2-FGF-21-mut) luciferase reporter gene carriers were constructed.The relevant plasmids 〔hsa-miR-577mimics, hsa-miR-583 mimics or miR negative control (miR-NC)〕 and luciferase reporter gene carrier (wild type or mutant ) were co-transfected into 293T cells.The luciferase reporter system was used to detect the luciferase activity.The effects of has-miR-577 and has-miR-583 on the expression of FGF-21 were observed. Results The double enzyme electrophoresis and sequencing results showed that the gene fragment size and sequences of the wild type (psiCHECK2-FGF-21) and mutant (psiCHECK2-FGF-21-mut) carriers met expectations of the experiment. The luciferase assays revealed that has-miR-577 and has-miR-583 significantly diminished luciferase activity from the reporter vector containing 3′UTR of FGF-21 (PFGF-21-mut).Conclusion FGF-21 gene can be targeted by has-miR-577 and has-miR-583.  
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