Functional Verification Of The SsDHN Promoter Of S.salsa (L) Pall

The promoter plays an important role in gene expression and regulation.Our team has cloned the Ss DHN gene and obtained its promoter sequence.basing on predicted cis-acting elements we constructed the deletion fragment of this promoter.In this study,using the deleted fragments of Ss DHN promoter,We transformed them to the S.salsa?L?Pall by itransient expression technology,and transformed them to the toarabidopsis thaliana by Agrobacterium-mediated transformation technology.Then we sprayed the transgenic plants with salt and corresponding hormones,and we verified the promoter's function by GUS histochemical assays and real-time fluorescent quantitative PCR.The results of the study are as following:1.Sequence analysis of 851 bp Ss DHN promoter by bioinformatics online software showed the promoter contained not only the characteristic core elements of promoters such as TATA-box and CAAT-box,but also contained cis-acting elements that respond the hormone such as ABA,SA,MEJA,etc.2.In order to verify the function of the Ss DHN promoter,We transformed the different deletion fragments of Ss DHN promoter into the S.salsa?L?Pall by transient expressiontechnology,and sprayed the transgenic plants with 300 mmol�L^-1 Na Cl,100?mol�L^-1 ABA?SA?MEJA.GUS histochemical staining showed that full-length promoter fragment DM1 was induced by Na Cl,ABA,SA and MEJA,the deleted fragment of DM2 was induced by ABA and SA,the deleted fragments of DM3?DM4 were only induced by ABA.3.We transformed the deletion fragments of Ss DHN promoters into the arabidopsis thaliana by Agrobacterium-mediated transformation technology We obtained transgenic arabidopsis with DM1?DM2,and each transgene obtained 3 transgenic lines.u Using PCR,Western blot and GFP gene expression technology,we idetificate of the transgenic plants.The results of PCR showed that the bands were same as the target genes.Laser confocal microscopy showed the GFP gene was expressed.Western blot results showed that GFP protein was successfully expressed in arabidopsis thaliana.All the results indicated that exogenous genes DM1 and DM2 were successfully transferred into arabidopsis thaliana.We gained 3 plants of DM1 and DM2.4.In order to further verify the function of the Ss DHN promoter,300 mmol�L^-1 Na Cl and 100 ?mol�L^-1 ABA?SA?MEJA were treated to transgenic Arabidopsis thalianas.GUShistochemical staining and Real-time fluorescent quantitative PCR showed that DM1 with full-length promoter fragment was induced by Na Cl,ABA,SA,and MEJA,DM2 with deleted fragments was only induced by ABA and SA.