稳定转染hSOD1-G93A基因的肌萎缩侧索硬化细胞模型的建立

发布时间：2018-04-28 09:50

本文选题：hSOD-GA基因 + 肌萎缩侧索硬化　；参考：《中国老年学杂志》2017年03期

【摘要】：目的建立稳定转染h SOD1-G93A真核表达载体肌萎缩侧索硬化细胞模型。方法进行NSC-34细胞的培养,利用脂质体方法把已经构建好的pc DNA3.1(-)、h SOD1-WT-pc DNA3.1(-)和h SOD1-G93A-pc DNA3.1(-)质粒转染至NSC-34细胞系中,再通过G418筛选建立稳定表达h SOD1基因的NSC-34细胞系;以RT-PCR法鉴定转染后h SOD1在NSC34细胞系中的表达。结果转染h SOD1-G93A-pc DNA3.1(-)质粒的细胞与正常NSC-34细胞相比,胞体变圆,突起减少,且轴突变短。而转染pc DNA3.1(-)和h SOD1-WT-pc DNA3.1(-)质粒的细胞较正常NSC-34细胞无明显变化。RT-PCR法鉴定结果显示,转染h SOD1-WT-pc DNA3.1(-)和h SOD1-G93A-pc DNA3.1(-)质粒的细胞中h SOD1基因表达为阳性,而正常NSC-34细胞和转染pc DNA3.1(-)质粒的对照组为阴性。结论成功获得可稳定表达h SOD1-G93A基因的肌萎缩侧索硬化细胞模型。
[Abstract]:Objective to establish an amyotrophic lateral sclerosing cell model with stable transfection of h SOD1-G93A eukaryotic expression vector. Methods NSC-34 cells were cultured and transfected into NSC-34 cell line by liposome method. The constructed plasmids were transfected into NSC-34 cell line. The NSC-34 cell lines expressing h SOD1 gene stably were established by G418 screening. The expression of h SOD1 in NSC34 cell line was identified by RT-PCR assay. Results compared with normal NSC-34 cells, the cells transfected with h SOD1-G93A-pc DNA3.1 ~ (-1) plasmid had round cell bodies, fewer processes and shorter axons. The results of RT-PCR analysis showed that the expression of h SOD1 gene was positive in the cells transfected with the plasmid pcDNA3.1 ~ (-1) and h SOD1-WT-pc DNA3.1 ~ (-1). The results of RT-PCR showed that the expression of h SOD1 gene was positive in the cells transfected with the plasmid "h SOD1-WT-pc DNA3.1" and "h SOD1-G93A-pc DNA3.1 ~ (-1)". The normal NSC-34 cells and the control group transfected with pcDNA3.1 ~ (-) plasmid were negative. Conclusion the model of amyotrophic lateral sclerosis cells which can stably express h SOD1-G93A gene was successfully obtained.
【作者单位】：临沂市人民医院神经内科;临沂卫生学校;临沂市人民医院消化内科;临沂市人民医院中心实验室;
【分类号】：R741.02

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