Abstract
SummaryBidirectional green tissue‐specific promoters have important application prospects in genetic engineering and crop genetic improvement. However, there is no report on the application of them, mainly due to undiscovered natural bidirectional green tissue‐specific promoters and the lack of a comprehensive approach for the synthesis of these promoters. In order to compensate for this vacancy, the present study reports a novel strategy for the expression regulatory sequence selection and the bidirectional green tissue‐specific synthetic promoter construction. Based on this strategy, seven promoters were synthesized and introduced into rice by agrobacterium‐mediated transformation. The functional identification of these synthetic promoters was performed by the expression pattern of GFP and GUS reporter genes in two reverse directions in transgenic rice. The results indicated that all the synthetic promoters possessed bidirectional expression activities in transgenic rice, and four synthetic promoters (BiGSSP2, BiGSSP3, BiGSSP6, BiGSSP7) showed highly bidirectional expression efficiencies specifically in green tissues (leaf, sheath, panicle, stem), which could be widely applied to agricultural biotechnology. Our study provided a feasible strategy for the construction of synthetic promoters, and we successfully created four bidirectional green tissue‐specific synthetic promoters. It is the first report on bidirectional green tissue‐specific promoters that could be efficiently applied in genetic engineering.
Highlights
For the purpose of screening expected promoters, fusion promoters are usually constructed by appending large quantities of cis-regulatory elements or random sequences to the core promoter, and these fusion promoters are subsequently transformed into different bacteria or fungi to detect the expression patterns and abundance of the reporter gene (Jakob Vang et al, 2014; Sohoni et al, 2014)
We provided a feasible strategy for the construction of bidirectional tissue-specific synthetic promoter, and we successfully constructed and identified four bidirectional green tissue-specific synthetic promoters, which is the first report on bidirectional green tissue-specific promoters and provided significant promoters resources for genetic engineering
By means of GUS and GFP assays of the transgenic plants, explicit bidirectional and green tissue-specific expression pattern can be observed in all synthetic promoters constructed from the result of GUS histochemical stain and GFP fluorescence intensity, except the expression of GUS in endosperm of BiGSSP4 (Figure 2)
Summary
The morphogenesis, development and environmental adaptation of plants are closely related to the spatio-temporal expression of specific genes (Chen et al, 2010b; Mcelroy et al, 1990; Zhu et al, 2011). For the purpose of screening expected promoters, fusion promoters are usually constructed by appending large quantities of cis-regulatory elements or random sequences to the core promoter, and these fusion promoters are subsequently transformed into different bacteria or fungi to detect the expression patterns and abundance of the reporter gene (Jakob Vang et al, 2014; Sohoni et al, 2014). This strategy is not suitable for plant synthetic promoters because of the long transformation cycle and the vast identification procedures. The results showed that the reporter genes expressed after suffering from pathogens, salicylic acid, ethylene and jasmonate acid treatment
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