沟叶结缕草ZmPSY基因启动子对拟南芥的转化及功能分析

Functional Analysis of ZmPSY Gene Promoter from Zoysia matrella Using Transgenic Arabidopsis

利用染色体步移法克隆得到沟叶结缕草(Zoysia matrella)ZmPSY基因的启动子序列。将ZmPSY起始密码子上游的1512bp的启动子序列与GUS基因融合,构建了ZmPSYpro::GUS载体,并利用农杆菌介导的方法转化拟南芥。经潮霉素筛选和PCR鉴定,共获得8个转基因株系。GUS染色结果表明,ZmPSY基因启动子能够驱动GUS基因在拟南芥中表达,且集中在茎叶部位。利用荧光定量技术(qRT-PCR)分析了外施激素和非生物胁迫处理下GUS基因的表达量,结果表明:5μM ABA可诱导GUS表达增强,10μM MeJA和黑暗处理抑制GUS表达,而100μM GA3处理无显著变化。研究成功克隆ZmPSY基因的启动子序列,并证明ZmPSY基因的表达可受ABA、MeJA和黑暗处理的调控,为深入研究ZmPSY基因的转录调控提供了依据。

The promoter of ZmPSY gene was cloned from Zoysia matrella using genome walking method.The 1512 bp upstream sequence in front of the initiation codon of ZmPSY was infused with theβ-glucuronidase（GUS）reporter gene.Then transgenic Arabidopsis was generated using agrobacterium-mediated transformation.After homomycin screening and PCR verification,8 lines expressing ZmPSYpro：：GUS were finally selected and used in the following experiments.The results of GUS staining in transgenic plants showed that the ZmPSYpromoter could drive GUSgene expressing in Arabidopsis with the highest abundance in leaves.The qRT-PCR determination indicated that ZmPSY could be induced by exogenous5μM ABA,but suppressed by 10μM MeJA or dark treatment,respectively.However,exogenous 100μM GA3 showed no obvious effect.The study paved the way for further study of the transcriptional regulation mechanism and function of ZmPSY.