解放军医学院学报
解放軍醫學院學報
해방군의학원학보
Academic Journal of Chinese Pla Medical School
2013年
4期
382-385
,共4页
hHCN4基因%重组腺病毒%骨髓间充质干细胞%生物起搏
hHCN4基因%重組腺病毒%骨髓間充質榦細胞%生物起搏
hHCN4기인%중조선병독%골수간충질간세포%생물기박
hHCN4 gene%recombinant adenovirus%bone marrow mesenchymal stem cells%biological pacemaker
目的构建重组腺病毒载体Ad-hHCN4-IRES/eGFP在体外转染大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs),检测目的基因在BMSCs中mRNA和蛋白水平是否有表达.方法采用贴壁培养法分离纯化大鼠BMSCs;将携带hHCN4基因的重组腺病毒转染大鼠BMSCs,用荧光显微镜观察转染后BMSCs绿色荧光蛋白的表达,用RT-PCR和Western Blot法检测转染后hHCN4基因mRNA和蛋白水平的表达情况.结果 BMSCs成功贴壁分离传代.转染目的基因24 h后荧光显微镜下即观察到绿色荧光蛋白的表达;RT-PCR法检测到有hHCN4 mRNA的表达,Western blot法检测到有其蛋白水平的表达.结论重组腺病毒载体转染的BMSCs能够表达hHCN4基因.
目的構建重組腺病毒載體Ad-hHCN4-IRES/eGFP在體外轉染大鼠骨髓間充質榦細胞(bone marrow mesenchymal stem cells,BMSCs),檢測目的基因在BMSCs中mRNA和蛋白水平是否有錶達.方法採用貼壁培養法分離純化大鼠BMSCs;將攜帶hHCN4基因的重組腺病毒轉染大鼠BMSCs,用熒光顯微鏡觀察轉染後BMSCs綠色熒光蛋白的錶達,用RT-PCR和Western Blot法檢測轉染後hHCN4基因mRNA和蛋白水平的錶達情況.結果 BMSCs成功貼壁分離傳代.轉染目的基因24 h後熒光顯微鏡下即觀察到綠色熒光蛋白的錶達;RT-PCR法檢測到有hHCN4 mRNA的錶達,Western blot法檢測到有其蛋白水平的錶達.結論重組腺病毒載體轉染的BMSCs能夠錶達hHCN4基因.
목적구건중조선병독재체Ad-hHCN4-IRES/eGFP재체외전염대서골수간충질간세포(bone marrow mesenchymal stem cells,BMSCs),검측목적기인재BMSCs중mRNA화단백수평시부유표체.방법채용첩벽배양법분리순화대서BMSCs;장휴대hHCN4기인적중조선병독전염대서BMSCs,용형광현미경관찰전염후BMSCs록색형광단백적표체,용RT-PCR화Western Blot법검측전염후hHCN4기인mRNA화단백수평적표체정황.결과 BMSCs성공첩벽분리전대.전염목적기인24 h후형광현미경하즉관찰도록색형광단백적표체;RT-PCR법검측도유hHCN4 mRNA적표체,Western blot법검측도유기단백수평적표체.결론중조선병독재체전염적BMSCs능구표체hHCN4기인.
Objective To transfect the rat bone marrow mesenchymal stem cells (BMSCs) with recombinant adenovirus vector Ad-hHCN4-IRES/eGFP and detect the expression of target gene in hHCN4 mRNA and protein of BMSCs. Methods Rat BMSCs were isolated and purified by adherent culture and transfected with recombinant adenovirus vector containing human HCN4 gene. Expressions of green fluorescent protein and transfected hHCN4 mRNA and protein in transfected rat BMSCs were detected by fluorescence microscopy, RT-PCR and Western blot, respectively. Results The BMSCs were successfully isolated after adherent culture. Expressions of green fluorescent protein and transfected hHCN4 mRNA and protein in transfected rat BMSCs could be detected by fluorescence microscopy, PCR and Western blot, respectively. Conclusion Recombinant adenovirus vector-transfected BMSC can express the hHCN4 gene.
