Abstract
An effector–reporter system is a powerful tool used to study cellular signal transduction, but this technique has been traditionally used in protoplasts. A similar system to study cellular signal transduction in fruits has not yet been established. In this study, we aimed to establish an effector–reporter system for strawberry fruit, a model nonclimacteric fruit. We first investigated the characteristics of transient gene expression in strawberry fruits and found marked variation in gene expression levels among individual fruits, and this variation has complicated the establishment of a technical system. To overcome this difficulty, we investigated a sampling strategy based on a statistical analysis of the activity pattern of four different reporters (GUS, GFP, FLuc, and RLuc) among individual fruits and combinations of pairs of reporters (GUS/GFP and RLuc/FLuc). Based on an optimized sampling strategy, we finally established a step-by step protocol for the effector/reporter assay. Using FaMYB10 and FaWRKY71 as the effectors and GUS driven by the FaCHS promoter as the reporter, we demonstrated that this effector/reporter system was practical and reliable. This effector/reporter technique will contribute to an in-depth exploration of the signaling mechanism for the regulation of strawberry fruit ripening.
Highlights
Fruit ripening is a complex process that involves dramatic changes in various physiological and biochemical processes, such as color, sugar, acid, aroma, and textureassociated metabolisms[1,2,3,4,5]
Molecular studies have identified genes implicated in the regulation of fruit development and ripening, and these mainly include structural genes, such as chalcone synthase (CHS), chalcone isomerase (CHI), dihydroflavonol 4-reductase (DFR), flavanone 3-hydroxylase (F3H), flavonoid 3′-hydroxylase (F3′H), anthocyanidin synthase (ANS), UDP glucose-flavonoid 3-O-glucosyl transferase (UFGT), sucrose phosphate synthetase, and sucrose synthetase (SS), which are directly involved in physiological and biochemical metabolism
With GUS driven by the FaCHS promoter as the reporter and FaMYB10 as the effector, we investigated the response of the GUS reporter to FaMYB10 and to abscisic acid (ABA) and mannitol treatment
Summary
Fruit ripening is a complex process that involves dramatic changes in various physiological and biochemical processes, such as color, sugar, acid, aroma, and textureassociated metabolisms[1,2,3,4,5]. Based on their physiological ripening characteristics, fleshy fruits can be categorized into two major groups, climacteric and nonclimacteric fruits[6,7,8,9]. Less is known about the specific targets of TFs and the response of structural genes to the internal/external cues
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