摘要： 为进一步验证青花菜 BoPGIPl 基因的功能,通过RT-PCR方法克隆得到青花菜 BoPGIP1 基因完整的编码区序列,将其克隆到原核表达载体pET28a(+)上,转化大肠杆菌BL21(DE3),获得高效表达 BoPGIP1 基因的重组大肠杆菌BL21(pET28a- BoPGIP1),重组菌经IPTG诱导表达以及SDS-PAGE电泳检测,结果表明,在37 kDa处有一条特异表达的蛋白质条带,与预期的目的产物大小一致;同时对重组大肠杆菌BL21(pET28a- BoPGIP1)的抗逆性进行分析,结果表明,BL21(pET28a- BoPGIP1)对NaCl(0.4 mol/L)、NaHCO₃(0.2 mol/L)和PEG6000(20%)的抗性明显高于对照菌株BL21(pET28a),该抗性的证明为 BoPGIP1 基因在植物抗逆基因工程中的应用提供了理论依据。

关键词: 青花菜, BoPGIP1, 原核表达, 抗性分析

Abstract: In order to further verify the broccoli BopGIPI genes function, the complete coding sequence of BoPGIP1 (993 bp) for protein expression was amplified by RT-PCR with gene specific primers designed according to multiple clone sites in prokaryotic expression vector pET28a(+) and BoPGIP1 which was obtained in author's previous experiment.The gene was cloned into pET28a(+) vector, and transferred into E.coli strain BL21 (DE3) for heterologous expression after induced by IPTG.SDS-PAGE was used to analyze expression of the target protein.The result showed that there was a specific band at about 37 kDa in size, which was identical with the expected molecular weight of the recombinant protein.Meanwhile, stress tolerance of E.coli BL21 expressing BoPGIP1 gene was analyzed.The results indicated that E.coli line BL21(pET28a- BoPGIP1) had more obvious tolerance to salt, alkali and dry stress than E.coli BL21 (pET28a), these provide preliminary information for the application of BoPGIP1 gene in plant gene engineering.

Key words: Broccoli, BoPGIP1, Prokaryotic expression, Stress resistance

中图分类号: 

• Q78

• S634.03