| 慢病毒介导SOX9基因转染骨髓间充质细胞中的基因表达 |
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Received:August 25, 2012
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| 作者 | Author | 单位 | Unit | E-Mail |
| 白洁玉 |
BAI Jie-yu |
山西医科大学第二医院骨科, 山西 太原 030001 |
Department of Orthopaedics, the Second Hospital of Shanxi Medical University, Taiyuan 030001, Shanxi, China |
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| 梁大川 |
LIANG Da-chuan |
山西医科大学第二医院骨科, 山西 太原 030001 |
Department of Orthopaedics, the Second Hospital of Shanxi Medical University, Taiyuan 030001, Shanxi, China |
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| 程鹏 |
CHENG Peng |
山西医科大学第二医院骨科, 山西 太原 030001 |
Department of Orthopaedics, the Second Hospital of Shanxi Medical University, Taiyuan 030001, Shanxi, China |
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| 杨自权 |
YANG Zi-quan |
山西医科大学第二医院骨科, 山西 太原 030001 |
Department of Orthopaedics, the Second Hospital of Shanxi Medical University, Taiyuan 030001, Shanxi, China |
yzqonline@126.com |
| 卫小春 |
WEI Xiao-chun |
山西医科大学第二医院骨科, 山西 太原 030001 |
Department of Orthopaedics, the Second Hospital of Shanxi Medical University, Taiyuan 030001, Shanxi, China |
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| 期刊信息：《中国骨伤》2013年26卷，第7期，第597-600页 |
| DOI：10.3969/j.issn.1003-0034.2013.07.016 |
| 基金项目:国际科技合作项目(编号:2010DFA32450);国家自然科学基金资助项目(编号:30973048);国家人事部及山西省人事厅留学回国人员科技活动择优资助项目;山西省留学基金项目(编号:107);山西省自然科学基金资助项目(编号:2010011050-6) |
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目的: 构建携带小鼠SOX9基因的慢病毒载体,体外转染小鼠骨髓间充质细胞,观察小鼠SOX9基因在小鼠间充质细胞中的表达。
方法: 从含有小鼠SOX9基因的质粒提取总RNA,利用RT-PCR方法扩增目的基因。连接目的基因与经Age-I酶切线性化的慢病毒载体,转化感受态的大肠杆菌对质粒进行扩增,筛出阳性转化子,经293T细胞包装,收集病毒后,通过基因测序和限制性核酸内切酶酶切的方法对质粒进行鉴定。Lenti-SOX9-EGFP体外转染小鼠骨髓间充质细胞,利用倒置荧光显微镜观察转染是否成功,并通过流式细胞仪测定转染效率。同时利用RT-PCR和Western Blot检测小鼠SOX9基因的表达。
结果: 成功构建了携带SOX9基因慢病毒载体,Lenti-SOX9-EGFP能高效转染小鼠骨髓间充质细胞。RT-PCR和Western Blot检测显示经SOX9基因转染的小鼠骨髓间充质细胞表达目的基因产物。
结论: 利用慢病毒介导SOX9基因成功地转染小鼠骨髓间充质细胞,而且SOX9基因在小鼠骨髓间充质细胞中得到表达,这为SOX9修复软骨损伤的进一步研究奠定了基础。 |
| [关键词]:骨关节炎 细胞介导免疫 慢病毒属 SOX9基因 骨髓间充质细胞 |
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| Gene expression of bone mesenchymal stem cells transduced by the lentiviral vector of SOX9 gene |
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Abstract:
Objective: To construct one lentiviral vector containing mouse SRY-related high mobility group-box gene 9 (SOX9) and transfect the murine bone mesenchymal stem cells (mBMSCs) in vitro and observe the expression of target gene.
Methods: RNA from the vectors containing mouse SOX9 gene were extracted and SOX9 genes were amplified by reverse transcription-Polymerase Chain Reaction (RT-PCR). The SOX9 genes were connected into lentiviral vectors pGC-FU. Then pGC-FU-SOX9 transduced into 293T cells to produce recombinant lentivirus called as Lenti-SOX9-EGFP. mBMSCs were transfected. The expression of target gene was detected by immunofluorescence,RT-PCR and Western Blot.
Results: Lenti-SOX9-EGFP was recombined successfully and transduced efficiently into mBMSCs. The expression of SOX9 gene was confirmed by RT-PCR and Western Blot.
Conclusion: Lentiviral vector of mouse SOX9 gene can transfect successfully into mBMSCs. Meanwhile,SOX9 gene may be expressed in mBMSCs. This will provide the target cells for the following study about SOX9 gene repairing cartilage injury. |
| KEYWORDS:Osteoarthritis Cell-mediated immunity Lentivirus SOX9 gene Bone mesenchymal stem cells |
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| 引用本文，请按以下格式著录参考文献： |
| 中文格式: | 白洁玉,梁大川,程鹏,杨自权,卫小春.慢病毒介导SOX9基因转染骨髓间充质细胞中的基因表达[J].中国骨伤,2013,26(7):597~600 |
| 英文格式: | BAI Jie-yu,LIANG Da-chuan,CHENG Peng,YANG Zi-quan,WEI Xiao-chun.Gene expression of bone mesenchymal stem cells transduced by the lentiviral vector of SOX9 gene[J].zhongguo gu shang / China J Orthop Trauma ,2013,26(7):597~600 |
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