首页 | 本学科首页   官方微博 | 高级检索  
     

结缔组织生长因子基因RNA干扰复制缺陷型腺病毒载体的构建及鉴定
引用本文:郝春秋,彭梅娟,谢玉梅,周云,魏欣,马力,王素娜,李瑞娟,张岩,白雪帆,贾战生. 结缔组织生长因子基因RNA干扰复制缺陷型腺病毒载体的构建及鉴定[J]. 生物技术通讯, 2013, 0(1): 57-60
作者姓名:郝春秋  彭梅娟  谢玉梅  周云  魏欣  马力  王素娜  李瑞娟  张岩  白雪帆  贾战生
作者单位:第四军医大学 唐都医院感染病诊疗中心,陕西 西安 710038
基金项目:国家自然科学基金(30571658)
摘    要:目的:构建能介导结缔组织生长因子(CTGF)基因RNA干扰的复制缺陷型腺病毒表达载体。方法:以大鼠CTGF基因为靶序列,设计并合成含编码短发夹RNA序列的寡核苷酸,构建腺病毒穿梭质粒P-shuttle-CTGF,酶切及测序分析正确后,与腺病毒骨架质粒pAdEasy-1共转染AD-293细胞,进行病毒包装,得到腺病毒载体Ad.H1-CTGF,用该载体感染HSC-T6细胞,观察其对CTGF基因表达抑制的效果。结果:构建的腺病毒穿梭质粒p-shuttle-CTGF经酶切、测序分析证实正确;包装的病毒载体滴度为4×1010PFU/mL,感染HSC-T6细胞后,Western印迹证实CTGF表达显著减少。结论:构建的腺病毒载体Ad.H1-CTGF可有效抑制HSC-T6中CTGF的表达,为抗纤维化研究提供了有力的工具。

关 键 词:结缔组织生长因子  RNA干扰  腺病毒载体

Construction and Identification of Replication-Deficient Adenovirus Vector for RNA Interference with CTGF Gene
HAO Chun-Qiu,PENG Mei-Juan,XIE Yu-Mei,ZHOU Yun,WEI Xin,MA Li,WANG Su-Na,LI Rui-Juan,ZHANG Yan,BAI Xue-Fan,JIA Zhan-Sheng. Construction and Identification of Replication-Deficient Adenovirus Vector for RNA Interference with CTGF Gene[J]. Letters in Biotechnology, 2013, 0(1): 57-60
Authors:HAO Chun-Qiu  PENG Mei-Juan  XIE Yu-Mei  ZHOU Yun  WEI Xin  MA Li  WANG Su-Na  LI Rui-Juan  ZHANG Yan  BAI Xue-Fan  JIA Zhan-Sheng
Affiliation:Center of Infectious Diseases of Chinese PLA,Affiliated Tangdu Hospital,Fourth Military Medical University,Xi’an 710038,China
Abstract:Objective: To construct a replication-deficient recombinant adenovirus vector expressing small interfer- ence RNA(siRNA) against the connective tissue growth factor(CTGF) gene. Methods: The siRNA sequence target- ing CTGF mRNA was synthesized and shuttle plasmid p-shuttle-CTGF was constructed, which was co-transfected into AD-293 cells with the bone plasmid pAdEasy-1 to obtain the homologous recombination adenovirus vector Ad.H1-CTGF. The concentration of CTGF protein in HSC-T6 cells infected with adenovirus vector Ad.H1-CTGF was determined by Wetsern blot. Results: The p-shuttle-CTGF was identified by endonuclease and sequencing. The adenovirus vector Ad.H1-CTGF was successfully constructed with the titre of 4×1010 PFU/mL by purification. The concentration of CTGF protein in HSC-T6 ceils was decreased observably after Ad.H1-CTGF infected by Western blot. Conclusion: The constructed Ad.H1-CTGF can effectively inhit the expression of CTGF in HSC-T6 ceils, which provided a powerful tool for anti-fibrosis study.
Keywords:connective tissue growth factor  RNA interference  adenovirus vector
本文献已被 CNKI 维普 万方数据 等数据库收录!
设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号