[1]孟凡萍,杨刚毅,李钶,等.腺病毒载体介导的siRNA对3T3-L1细胞脂联素表达的影响[J].第三军医大学学报,2008,30(14):1312-1315.
 MENG Fan-ping,YANG Gang-yi,LI Ke,et al.Effects of recombinant adenovirus-mediated siRNA on adiponectin expression in 3T3-L1 cells[J].J Third Mil Med Univ,2008,30(14):1312-1315.
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腺病毒载体介导的siRNA对3T3-L1细胞脂联素表达的影响(/HTML )
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
30卷
期数:
2008年第14期
页码:
1312-1315
栏目:
论著
出版日期:
2008-07-30

文章信息/Info

Title:
Effects of recombinant adenovirus-mediated siRNA on adiponectin expression in 3T3-L1 cells
作者:
孟凡萍杨刚毅李钶李伶齐晓亚刘建雷李荣
重庆医科大学:医学检验系临床生物化学教研室,临床检验诊断学省部共建教育部重点实验室,附属第二医院内分泌科
Author(s):
MENG Fan-ping YANG Gang-yi LI Ke LI Ling QI Xiao-ya LIU Jian-lei LI Rong
Department of Clinical Biochemistry,Key Laboratory of Laboratory Medical Diagnostics of Ministry of Education, Faculty of Laboratory Medicine,  Chongqing 400016; Department of Endocrinology, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, China
关键词:
RNA干扰腺病毒载体脂联素3T3-L1细胞
Keywords:
RNA interference adenovirus vector adiponectin 3T3-L1 cell
分类号:
R329.25;R394-33;R394.2
文献标志码:
A
摘要:
目的  构建携带小鼠脂联素(Acrp30)siRNA腺病毒载体,并检测其对小鼠脂肪细胞Acrp30表达的影响。 方法  首先用KpnⅠ+NotⅠ双酶切本课题组前期构建的质粒pSilencer1.0-U6-Acrp30-1和pSilencer1.0-U6-Acrp30-3,得到目的片段U6-Acrp30-1和U6-Acrp30-3,并将目的片段亚克隆入穿梭质粒pShuttle,用PmeⅠ线性化穿梭质粒pShuttle-U6-Acrp30-1和pShuttle-U6-Acrp30-3,并与pAdeasy-1在BJ5183内同源重组,筛选、鉴定、测序后,在XL10-Gold中扩增重组腺病毒质粒,最后在293细胞内包装扩增为重组腺病毒Ad-Acrp30-1和Ad-Acrp30-3。用此重组腺病毒感染3T3-L1脂肪细胞,用RT-PCR和ELISA检测其Acrp30 mRNA和蛋白表达。  结果  设计并构建了小鼠Acrp30 基因特异性siRNA腺病毒载体,该载体感染脂肪细胞后,能显著抑制其Acrp30 mRNA和蛋白表达。  结论  构建的Acrp30 基因特异性siRNA腺病毒载体能有效的抑制脂联素基因在3T3-L1脂肪细胞中的表达。
Abstract:
Objective    To construct the RNA interfered adenovirus expression vector specific for Adiponectin (Acrp30) gene and observe its effect on the Acrp30 expression in 3T3-L1 cells.     Methods    Firstly, fragments U6-Acrp30-1 and U6-Acrp30-3 were obtained by using KpnⅠ+NotⅠ to digest the plasmids pSilencer1.0-U6-Acrp30-1 and pSilencer1.0- U6-Acrp30-3.  Two adenovirus vector plasmids, Ad-Acrp30-1 and Ad-Acrp30-3 were constructed according to a two-step transformation protocol. Two newly constructed plasmids were transfected into 293 packaging cells to produce adenovirus, which were further multiplied and purified. Then 3T3-L1 cells were infected with the 2 recombinant adenoviruses respectively. The mRNA expression and protein levels of Acrp30 in these cells were evaluated by semi-quantitative RT-PCR and ELISA.     Results    Two recombinant adenoviral plasmids Ad-Acrp30-1 and Ad-Acrp30-3 were successfully constructed. They-remarkably down-regulated the expression of Acrp30 at both the mRNA and protein levels in transfected 3T3-L1 cells.     Conclusion    The siRNA eukaryotic expression vectors against Acrp30 mRNA effectively inhibit the expression of Acrp30 in 3T3-L1 adipocytes.

参考文献/References:

孟凡萍,杨刚毅,李钶,等.  腺病毒载体介导的siRNA对3T3-L1细胞脂联素表达的影响[J].第三军医大学学报,2008,30(14):1312-1315.

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更新日期/Last Update: 2008-07-17