大鼠GDNF及EDNRB基因共重组腺病毒的构建及在神经干细胞中的共表达
发布时间：2018-07-28 06:53
【摘要】： 目的:构建大鼠胶质细胞源性营养因子(GDNF)及内皮素B受体(EDNRB)基因共表达腺病毒并观察其在神经干细胞(NSC)中的表达。方法:利用Ad-Easy腺病毒表达系统构建GDNF及EDNRB基因共表达腺病毒PAd-GE,体外感染NSC后通过观察绿色荧光蛋白表达及RT-PCR、实时荧光定量PCR.流式细胞仪法鉴定外源基因的表达。结果:NSC经重组腺病毒PAd-GE感染后24h可观察到绿色荧光蛋白(GFP)表达,RT-PCR、实时荧光定量PCR、流式细胞仪法证实外源性GDNF及EDNRB基因均可在NSC中表达。结论:成功构建GDNF及EDNRB基因共表达重组腺病毒,证实了其在NSC的表达,为先天性巨结肠的联合基因治疗打下基础。
[Abstract]:Aim: to construct rat glial cell-derived nutrition factor (GDNF) and endothelin B receptor (EDNRB) gene co-expressing adenovirus and observe its expression in neural stem cell (NSC). Methods: GDNF and EDNRB co-expressed adenovirus PAd-GE was constructed by using Ad-Easy adenovirus expression system. The expression of green fluorescent protein and RT-PCR were observed after NSC infection in vitro. The expression of exogenous gene was identified by flow cytometry. Results Twenty-four hours after infection with recombinant adenovirus PAd-GE, the expression of green fluorescent protein (GFP) was observed, and real-time fluorescence quantitative PCR was detected. Flow cytometry confirmed that exogenous GDNF and EDNRB genes could be expressed in NSC. Conclusion: the recombinant adenovirus of GDNF and EDNRB gene was successfully constructed, which confirmed its expression in NSC and laid the foundation for combined gene therapy of Hirschsprung's disease.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2007
【分类号】：R656.9;R346
[Abstract]:Aim: to construct rat glial cell-derived nutrition factor (GDNF) and endothelin B receptor (EDNRB) gene co-expressing adenovirus and observe its expression in neural stem cell (NSC). Methods: GDNF and EDNRB co-expressed adenovirus PAd-GE was constructed by using Ad-Easy adenovirus expression system. The expression of green fluorescent protein and RT-PCR were observed after NSC infection in vitro. The expression of exogenous gene was identified by flow cytometry. Results Twenty-four hours after infection with recombinant adenovirus PAd-GE, the expression of green fluorescent protein (GFP) was observed, and real-time fluorescence quantitative PCR was detected. Flow cytometry confirmed that exogenous GDNF and EDNRB genes could be expressed in NSC. Conclusion: the recombinant adenovirus of GDNF and EDNRB gene was successfully constructed, which confirmed its expression in NSC and laid the foundation for combined gene therapy of Hirschsprung's disease.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2007
【分类号】：R656.9;R346
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