论文部分内容阅读
目的:构建Sonic Hedgehog(Shh)基因慢病毒载体,并进行鉴定。方法:构建携带Shh基因慢病毒表达质粒载体p LV.Ex2d.P/puro-EF1A-m Shh,RT-PCR反应和基因测序证实质粒载体构建成功。对载体进行包装,测定病毒滴度。将携带Shh基因慢病毒颗粒以感染复数(MOI=10)转导HT1080细胞,使用RT-PCR法检测转导细胞中Shh基因的表达。结果:成功构建携带Shh基因的慢病毒表达载体(Lenti-Shh),病毒滴度为2.9×10~7TU/m L。RTPCR检测结果证明Lenti-Shh转导细胞成功表达Shh基因。结论:成功构建小鼠Shh慢病毒载体系统,为基因增强型骨组织工程提供可靠的基因传递载体。
Objective: To construct Sonic Hedgehog (Shh) lentiviral vector and identify it. Methods: Construction of lentiviral vector containing lentivirus Shh gene p.Ex2d.P / puro-EF1A-m Shh, RT-PCR and gene sequencing confirmed that the plasmid vector was constructed successfully. The vector is packaged and the virus titer is determined. The Shh gene lentivirus particles were transduced into HT1080 cells at a multiplicity of infection (MOI = 10), and the Shh gene expression in the transduced cells was detected by RT-PCR. Results: The Lenti-Shh vector carrying Shh gene was successfully constructed and the virus titer was 2.9 × 10 ~ 7 TU / m L. RTPCR test results showed that Lenti-Shh transduced cells successfully expressed Shh gene. Conclusion: The mouse Shh lentiviral vector system was successfully constructed and provided a reliable gene delivery vector for gene-enhanced bone tissue engineering.