摘要
目的:构建转化生长因子β1(TGF-β1)表达载体,在骨髓间充质干细胞(BMSC)中表达。方法:以小鼠肺组织cDNA为模板,PCR扩增TGF-β1基因,并将其插入pCDH1-MCS1-EF1-copGFP载体质粒,转化感受态大肠杆菌DH5α,抽提质粒,经PCR和测序鉴定后转染BMSC,利用激光共聚焦显微镜和Western印迹对其表达进行检测。结果:经PCR及测序鉴定,构建入载体质粒的基因为TGF-β1基因,pCDH1-TGFβ1-EF1-copGFP重组质粒能在BMSC中表达。结论:构建了pCDH1-TGFβ1-EF1-copGFP重组质粒,且能表达于BMSC,为进一步研究TGF-β1影响间充质干细胞的生理功能奠定了基础。
Objective: To construct eukaryotic expression vector of the transforming growth factor β1 (TGF-β1), and express the fusion protein TGF-β1-copGFP in mouse marrow mesenehymal stem cells(BMSC). Methods: The full-length coding sequence of TGF-β1 was amplified from the eDNA of mouse lung sample by PCR, and was cloned into the Xba I/BstB I sites of the pCDH1-EF1-copGFP expression vector. After sequencing, the recombinant plasmid was transfected into BMSC and was observed by laser eonfocal microscopy and Western blot. Results: The results of PCR and sequencing showed that the recombinant plasmid pCDH1-TGFβ1-EF1-copGFP was constructed. What's more, the TGF-β1-eopGFP protein can expressed in BMSC. Conclusion: The recombinant plasmid pCDH1-TGFβ1-EF1-copGFP was constructed and it can express the protein TGF-β1 in BMSC, which provided a better understanding of the biological behavior of BMSC regulated by TGF-β1.
出处
《生物技术通讯》
CAS
2013年第6期797-800,共4页
Letters in Biotechnology
基金
国家自然科学基金(81170602)
关键词
转化生长因子Β1
慢病毒载体
骨髓间充质干细胞
transforming growth factor β1
lentiviral vector
bone marrow mesenchymal stem cells