摘要
目的 :构建能够用于骨缺血研究的能表达huVEGF12 1的重组腺病毒载体。方法 :将huVEGF12 1克隆至穿梭质粒 ,线性化后转染含骨架质粒的BJ5 183菌 ,构建重组腺病毒质粒。酶切及PCR法鉴定 ;将其线性化后转染 2 93细胞 ,通过观察绿色荧光监测病毒产生 ,免疫组化检测VEGF12 1表达。结果 :成功的将hu -VEGF12 1克隆至穿梭质粒pTrack -CMV ,正确构建了重组腺病毒质粒 ,感染 2 93细胞得到大量病毒。结论 :成功构建了huVEGF12 1重组腺病毒 ,为缺血性骨病的进一步研究奠定了基础。
Objective:To construct the adenoviral vector that expresses hu-VEGF 121 gene for further study of the gene therapy on the osteonecrosis.Methods: The hu-VEGF 121 gene was cloned to the shuttle plasmid Track-CMV, the shuttle plasmids were linearized with Pme1 and was transfected to BJ5183 cells which obtain adenoviral backbone vector. The candidate clones were further tested by nuclease digestion and PCR. The recombinant adenoviral plasmids were linearized and transfected to 293 cells, viral production were monitored by GFP(green fluorescent protein)expression. The expression of VEGF 121 was tested by immunohistochemistry. Results:We successfully cloned the hu-VEGF 121 to the shuttle plasmid Track-CMV. The correct recombinant adenoviral plasmid was constructed and recombinant adenovirus hu-VEGF 121 was obtained by transfecting 293 cells.Conclusion:We have constructed the recombinant adenovirus hu-VEGF 121 ,it will provides basis for the further study of gene therapy on the osteonecrosis.
出处
《西北国防医学杂志》
CAS
2005年第1期7-9,F003,共4页
Medical Journal of National Defending Forces in Northwest China
基金
重庆市科委应用基础项目基金资助项目 (2 0 0 3C0 5 3 )
