转化生长因子β1基因真核表达质粒的构建及其在小鼠骨髓间充质干细胞中的表达

摘要

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 mesenchymal stem cells(BMSC). Methods: The full-length coding sequence of TGF-β1 was amplified from the cDNA of mouse lung sample by PCR, and was cloned into the XbaⅠ/BstBⅠ sites of the pCDH1-EF1-copGFP expression vector. After sequencing, the recombi-nant plasmid was transfected into BMSC and was observed by laser confocal microscopy and Western blot. Re-sults: The results of PCR and sequencing showed that the recombinant plasmid pCDH1-TGFβ1-EF1-copGFP was constructed. What's more, the TGF-β1-copGFP 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.%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。