Characterization of a novel stilbene synthase promoter involved in pathogen- and stress-inducible expression from Chinese wild Vitis pseudoreticulata

Abstract

Stilbene synthase is a plant-specific polyketide synthase, and plays important roles in diverse metabolic processes. The genomic stilbene synthase gene was cloned from accession "Baihe-35-1" of Chinese wild Vitis pseudoreticulata, and a stilbene synthase of V. pseudoreticulata (VpSTS) transcripts expressed in the grape-powdery mildew interaction were determined by semi-quantitative RT-PCR. To monitor VpSTS expression in plant, the promoter region flanking the 5' VpSTS coding region was isolated from the genomic DNA of Chinese wild V. pseudoreticulata accession Baihe-35-1. Alignment of the VpSTS promoter sequence showed a 56.4% identity to Vitis vinifera. To identify the upstream region of the VpSTS gene required for promoter activity, a series of VpSTS promoter deletion derivatives was constructed. Each deletion construct was analyzed by Agrobacterium-mediated transient transformation in grapevine and tobacco leaves after infection by Uncinula necator and Alternaria alternata. In transiently transformed grapevine leaves, GUS activity was also determined after treatment with salicylic acid (SA) and 4 degrees C cold. Analysis of a series of 5' deletions of the VpSTS promoter in grapevine leaves indicated that the proximal 162 bp from the transcription initiation site was proved to be necessary for establishing both the constitutive and induced pattern of expression.

Promoters for Transgenic Horticultural Plants

“…It was detected that the proximal 162 bp from the transcription initiation site were enough to keep the specifi c transcription pattern. Alignment of the stilbene synthase promoter sequence from a PM-resistant Chinese wild V. pseudoreticulata showed only a 56.4 % identity to stilbene synthase promoter from a PM-susceptible cultivated grapevine V. vinifera (Xu et al 2010 ).…”

Expression and functional analysis of two genes encoding transcription factors, VpWRKY1 and VpWRKY2, isolated from Chinese wild Vitis pseudoreticulata

“…The same procedure was done on the fragment of VpWRKY2 from 24 to 323 bp to generate the VpWRKY2 silencing construct. Transient overexpression and silencing of VpWRKY1 and VpWRKY2 in 'Baihe-35-1' leaves was performed as described by Xu et al (2010). qRT-PCR was used to analyze expression of defense marker genes at 3 days post infiltration.…”

“…Agrobacterium strain GV3101 harboring recombinant plasmids was transformed in vitro into 6-week-old plantlet leaves of V. pseudoreticulata W. T. Wang 'Baihe-35-1' via Agrobacterium-mediated transient assay. GUS staining was performed at 3 dpi as described by Xu et al (2010).…”

Characterization of FeDREB1 promoter involved in cold- and drought-inducible expression from common buckwheat (Fagopyrum esculentum)

“…To generate the reporter plasmids for the Agrobacterium-mediated transient expression system, the GUS gene from pCAMBIA1391 was cloned into the pCAMBIA0390 vector to generate pC0390::GUS and used as a negative control (Xu et al, 2010). The CaMV 35S promoter from pBI121 was introduced into pC0390GUS to generate the positive control pC35S::GUS construct.…”