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目的 构建 β 神经生长因子 (β NGF)基因重组真核表达载体 ,为 β NGF基因治疗和临床实验诊断研究打下良好的基础。方法 将质粒PGEM β NGF进行酶切获得 β NGF基因片段 ,用T4连接酶将其插入真核表达载体PcDNA3;以T7、P2为引物进行PCR ,论证插入片段的方向 ;经测序和酶切对插入片段进行分析和进一步鉴定。结果 PCR产物琼脂糖电泳结果显示 :在预期位置有阳性条带 ;序列分析和酶切结果证实插入片段序列正确。结论 β NGF基因重组真核表达载体PcDNA3 β NGF构建成功 ,为进一步开展 β NGF基因治疗神经系统疾病和临床实验诊断奠定了基础。
Objective To construct a recombinant eukaryotic expression vector of β-NGF gene and lay a good foundation for gene therapy and clinical laboratory diagnosis of β-NGF. Methods The fragment of β NGF gene was digested with PGEM β NGF and inserted into eukaryotic expression vector pcDNA3 using T4 ligase. The PCR was carried out with T7 and P2 as the primers to confirm the orientation of the inserted fragment. After sequencing and digestion, Fragment for analysis and further identification. Results The results of agarose gel electrophoresis of PCR products showed that there was a positive band at the expected position. Sequence analysis and restriction analysis confirmed that the inserted fragment was correct. Conclusion The recombinant eukaryotic expression vector pcDNA3 β NGF of β-NGF gene was constructed successfully, which lays the foundation for the further development of β-NGF gene in the treatment of nervous system diseases and clinical laboratory diagnosis.