Abstract
Background: The non-expressor of pathogenesis related gene 1 (NPR1) protein is one of the key regulators in the systemic acquired resistance plant defence system. The cis-acting elements of its distal promoter gene are characterized by salicylic acid inducing elements such as the W-box, RAV1AAT and ASF1, accompanied with enhancer and silencer elements. This study was aimed to isolate and characterize the distal promoter sequence of the NPR1 gene (PD_CbNPR1) from the chili pepper (Capsicum annuum L.) genotype Berangkai, a local genotype known to produce large yields, but is susceptible to viral infection. Elucidating its sequence structure will open a broad range of possibilities to engineer the NPR1 gene expression which is important to improve chili pepper resistant. Methods: PCR-based cloning combined with a primer walking strategy was applied in this study. The BioEdit tool was used to edit the sequence and verify sequence integrity, while homology analysis was conducted with BLASTn searching. Identification of a cis-acting element was detected by PLACE. Results: Isolation of the complete distal promoter sequence of PD_CbNPR1 produced a fragment 5,950 bp in size. BLASTn search analysis indicated that PD_CbNPR1 sequence is highly conserved (99% homology) showing only a single nucleotide polymorphism (SNP) (base substitution) compared with its reference sequence. Analysis using PLACE tools successfully identified nine cis-acting elements containing a W-box, WLE1, RAV1AAT, TATA-box, CAAT-box, GARE and GT1 with multi repeats and diverse motives, as well as enhancer and silencer elements, which is characterized by a CCAAT-box and GAGAAATT pattern, respectively. Conclusion: The distal promoter of the NPR1 gene is highly conserved, showing only one SNP caused by one base substitution event.
Highlights
The non-expressor of pathogenesis related gene 1 (NPR1) protein is a main regulator in the systemic acquired resistance response of many plants
Over expression by modifying distal promoter in the W-box element of the OsNPR1 gene in rice could increase its resistance against Xanthomonas oryzae pv
Similar results were reported by Zhong et al (2015), who modified the RAV1 element on the GhNPR1 which increased Gladiolus hybridus resistance against Curvularia gladioli by up to 18.6 times compared to control
Summary
The non-expressor of pathogenesis related gene 1 (NPR1) protein is a main regulator in the systemic acquired resistance response of many plants. Similar results were reported by Zhong et al (2015), who modified the RAV1 element on the GhNPR1 which increased Gladiolus hybridus resistance against Curvularia gladioli by up to 18.6 times compared to control. Based on those studies, we expect more prospects of the NPR1 gene promoter in the improvement of plant resistance against many pathogens. This study was aimed to isolate and characterize the distal promoter sequence of the NPR1 gene ( PD_CbNPR1) from the chili pepper (Capsicum annuum L.) genotype Berangkai, a local genotype known to produce large yields, but is susceptible to viral infection. Analysis using PLACE tools successfully identified nine cis-acting elements containing a W-box, WLE1, RAV1AAT, TATA-box, CAAT-box, GARE and GT1 with multi repeats and diverse motives, as well as enhancer and version 2
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