马铃薯WRKY6基因的克隆、序列分析与原核表达研究

Cloning,sequence analysis and prokaryotic expression of the WRKY6 of potato

在高等植物中WRKY转录因子家族成员众多,它广泛参与植物对生物胁迫、非生物胁迫、生长发育和代谢过程的调控。本研究采用同源序列克隆的方法获得马铃薯WRKY6基因,序列分析表明该蛋白属于WRKY家族第3组成员,锌指结构为C-X7-C-X23-H-X-C。构建系统发育树结果表明它与拟南芥WRKY70亲缘关系较近,相似性达58%。然后将其完整编码区构建到大肠杆菌表达载体。在培养温度18℃条件下添加0.2 mmol/L IPTG(异丙基硫代半乳糖苷),诱导培养8 h可获得高表达融合蛋白。进一步实验获得特异性和纯度非常高的纯化蛋白。本研究为进一步确定该蛋白的体外活性及生物学功能奠定了坚实基础。

In higher plants,there are many WRKY transcription factors in WRKY family.They are widely involved in biotic stress;abiotic stress;growth;development and metabolism regulation.Cloning WRKY6 from potato by homology cloning.Analysis show that the WRKY6 belongs to the third group of WRKY family members,zinc-finger structure is C-X7-C-X23-H-X-C.Phylogeny results show WRKY6 is the most closest to AtWRKY70 with 58% similarity,By SDS-PAGE analysis,the best expression conditions of WRKY6 are that host bacteria-Escherichia coli（DE3） is growing for 8 hours under 18℃ adding 0.2 mmol/L IPTG（Isopropyl β-D-thiogalactoside）,purified protein of high specificity is obtained.This research lay a solid foundation to further study the foundation of WRKY6.