The Preliminary Study On The Effect Of NPR1Gene Expression On Plant Disease Resistance Signaling Pathways In Arabidopsis Pumila

NPR1gene is a positive regulator of salicylic acid-induced systemic acquired disease resistance （SAR）, it is located in upstream of salicylic acid （SA） and downstream of pathogenesis-related protein genes, and itresponses to salicylic acid （SA）. In this study, the experiment was conducted with Arabidopsis Pumila, theNPR1gene was cloned in Arabidopsis Pumila, sequencing and bioinformatics analyses indicated that: a110bp intron of NPR1gene was not cut when transcription. It is currently unclear whether this intron affects the expression of NPR1gene and its downstream resistance related genes. The plant recombinant plasmids ofpBI121:: Ap.NPR1and pCambia2300:: pUCCRNAiN₂N₃were constructed by PCR, enzyme digestion andother molecular biology methods. They were transfored by frozen into Agrobacterium tumefaciens GV3101wild type Arabidopsis thaliana and Arabidopsis Pumila were transformated by Agrobacterium using drop flowers and floral dip method. Transgenic plants were screened by kanamycin and identified by PCR, finallytransgenic postive plants were gained; the relative expression of NPR1gene and PR-1, PR-2, PR-3, PR-4, PR-5, PDF1.2gene induced by NPR1gene with different treatment （SA, MeJA） at different times weredetected by Real-time PCR. And then the plants were treated with salicylic acid, the enzyme activity ofPOD, CAT and SOD in transgenic plants were detrminated. The relative expression of NPR1, PR-1, PR-2, and PR-5genes in SAR pathway were detected by Real-time PCR. The results of showed that the relativeexpression of NPR1reached the maximum after treatment12h. the expression of PR-1, PR-2, PR-5genesreached the maximum after treatment60h,24h and12h respectively, and they were higher than wild typeArabidopsis thaliana. The mximum expression of NPR1, PR-1PR-2and PR-5genes in transgenic plantswere lower than those in wild type Arabidopsis thaliana. The results of relative expression of NPR1, PR-1, PR-2, and PR-5genes in ISR pathway showed that the expression of PDF1.2, PR-3and PR-5genesreached the maximum after treatment12h,60h and24h respectively, and they were higher than wild typeArabidopsis thaliana. The maximum expression of NPR1, PR-3, PR-4and PDF1.2genes in transgenicplants were lower than those in wild type Arabidopsis thaliana. The enzyme activity of POD and SOD inild type Arabidopsis Pumila were higher than that in wild type Arabidopsis thaliana, but the enzyme activeity of CAT in wild type Arabidopsis Pumila was lower than that in wild type Arabidopsis thaliana. However, the enzyme activity in transgenic plants changed little. In conclusion, the110bp intron of NPR1gene inwild type Arabidopsis Pumila is retained in transcription of mRNA compared with Arabidopsis thaliana, and the translation process was terminated prematurely by the change of ORF, so the expression of downstream genes （PR-1） can not be activated, but the present study showed that the expression of disease-related protein genes （PR-1） in wild type Arabidopsis Pumila is higher than shose in Arabidopsis thaliana in SAR pathway. We can putatively draw the conclusion that the resistance-related genes in downstream could be activated and induced not only by NPR1but also by other related genes. Plant disease resistance could be effectively improved by the expression of resistance-related genes and the H₂O₂which was accumulated by enhancing the enzyme activity of POD and SOD and inhibitting the enzyme activity of CAT. This study provides a theoretcal basis for the further research on resistance function of NPR1gene.