| SIK2 shRNA重组腺病毒载体构建及其在3T3-L1脂肪细胞的表达 |
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| 引用本文： | 杜静,杜玲,宴耀明,周薇,陆红,邹德学. SIK2 shRNA重组腺病毒载体构建及其在3T3-L1脂肪细胞的表达[J]. 中国实验诊断学, 2011, 15(10): 1614-1617 |
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| 作者姓名： | 杜静 杜玲 宴耀明 周薇 陆红 邹德学 |
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| 作者单位： | 1. 北京大学深圳医院,广东,深圳518036
2. 襄樊市中医院,湖北,襄樊441000 |
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| 基金项目： | 深圳市科技计划项目，留学回国人员科研启动基金 |
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| 摘 要： | 目的构建特异性小鼠盐诱导基因（Salt induced kinase2,SIK2）的shRNA腺病毒载体,干扰脂肪细胞中SIK2的表达。方法设计3对靶向SIK2的shRNA,插入穿梭质粒pENTR/U6载体,转染3T3-L1脂肪细胞,通过Real-timePCR方法筛选最佳沉默SIK2效果的穿梭质粒,经腺病毒同源重组后,重组小鼠SIK2shRNA腺病毒载体经脂质体转染HEK293A细胞,在细胞内包装获得腺病毒颗粒AdV-SIK2-shRNA,并进行大量扩增和纯化。纯化的腺病毒感染成熟的3T3-L1脂肪细胞,通过Real-time PCR及免疫印迹法检测鼠SIK2在3T3-L1脂肪细胞中的表达水平。结果纯化后的Ad-shRNA-SIK2重组腺病毒载体可特异下调成熟3T3-L1脂肪细胞SIK2 mRNA和SIK2蛋白的表达。结论干扰3T3-L1脂肪细胞SIK2表达的短发夹RNA重组腺病毒载体构建成功。
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| 关 键 词： | SIK2 腺病毒载体 短发夹RNA 3T3-L1脂肪细胞 |
Construction and expression of adenovirus vector of shRNA against SIK2 in in 3T3-L1 adipocytes |
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| DU Jing,DU Ling,YAN Yao-ming,et al.. Construction and expression of adenovirus vector of shRNA against SIK2 in in 3T3-L1 adipocytes[J]. Chinese Journal of Laboratory Diagnosis, 2011, 15(10): 1614-1617 |
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| Authors: | DU Jing DU Ling YAN Yao-ming et al. |
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| Affiliation: | DU Jing,DU Ling,YAN Yao-ming,et al.(Peking University Shenzhen Hospital,Shenzhen 518036,China) |
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| Abstract: | Objective To construct the recombinant adenovirus vector of short hairpin interfering RNA(shRNA) against SIK2(Salt induced kinase2) and explore the knocking-down expression of SIK2 in adipocytes.Methods Three short hairpin oligonucleotides and complementary strands were designed to target the consensus sequences of mouse SIK2.The BLOCK-iT RNAi system was used to construct shRNA.Real-time PCR was employed for screening the most effective pENTR/U6-shRNA-SIK2 entry vector.The entry vector was transfected into ... |
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| Keywords: | Mouse SIK2 Adenovirus shRNA 3T3-L1 adipocyte |
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