Abstract: 【Objective】In order to improve the structural stability of anthocyanins in wine grapes grown in the eastern foothills of the Helan Mountains, avoid rapid color withering during fruit ripening, and enhance the color stability of grapes and wine, this study selected one of the main wine grape varieties grown in the eastern foothills of the Helan Mountains‘Marselan’ (Vitis vinifera L.), to investigate the effects of shading at different color transformation stages on the expression of anthocyanin components and synthesis related genes in wine grape.【Methods】Using the wine grape‘Marselan’, as the test material, the top of the grape tree was shaded from the beginning of color transformation to the harvest period, and this treatment was T1 Sunshade the top of the grapevine from the complete color transition period to the harvest period, which is referred to as T2 treatment.Using trees that grow normally without shading as a blank control. The shading net adopted a black single-layer shading net with a shading rate of 50% and with a specification of 10 meters wide and 25 meters long. All three treatments are single experiments, and each treatment has three biological replicates. High-performance liquid chromatography (HPLC) was used to qualitatively and quantitatively analyze the content of anthocyanins, and the content and components of anthocyanins were studied. RNA transcriptome sequencing technology (RNA-seq) and real-time quantitative fluorescence PCR technology (qRT-PCR) were used to investigate the expression levels of genes related to anthocyanin synthesis.【Results】Sunshade can significantly reduce the soluble solids content during the ripening period of grape fruit, treatment at different stages can increase the titratable acid content of harvested fruits, and shading the tree during the complete color transition period can promote the accumulation of reducing sugar content and total anthocyanin content in the fruits. The 16 types of anthocyanins detected in each treatment were consistent, indicating that shading had no effect on the types of grape anthocyanins. At the same time, shading treatment increased the modification ratio of grape acylated anthocyanins, increased the stability of anthocyanin structure, and further improved the color stability of anthocyanins. After shading treatment, the content of delphinidin(Dp), cyanidin(Cy), petunidin(Pt) and peonidin (Pn)anthocyanins was significantly reduced, while shading treatment during the complete color transition period significantly increased the content of malvidin(Mv) anthocyanins, indicating that shading has different effects on anthocyanins at different stages. The synthesis of anthocyanins is influenced by both external environmental factors and the expression of key enzymes involved in the internal regulatory synthesis pathway.The study showed that the expression levels of functional genes related to anthocyanin synthesis were different after shading in different periods. During the initial stage of color transformation and shading during the harvesting period, the expression levels of CHI, DFR and OMT genes were upregulated compared to other treatments, with DFR and OMT reaching significant levels during the harvesting period. During the complete color transition period and shading during the harvesting period, the expression levels of PAL, CHS3, F3’H, F3’5’H, UFGT, GST4 and MYB90 genes were upregulated compared to other treatments during the harvesting period. qRT-PCR analysis revealed that among the 15 screened genes, the expression levels of CHI, F3H, F3’5’H, F3’H, DFR, LDOX, UFGT and OMT showed inconsistent trends with transcriptome sequencing in each treatment, while the changes in other genes were consistent with transcriptome sequencing results.【Conclusion】There is no significant difference in the effect of shading during the initial color transition period and the complete color transition period on fruit quality. The complete color transition period shading increased the transcription level of anthocyanin synthesis related genes, promoted anthocyanin synthesis, increased the total anthocyanin content of grapes, and significantly increased the content of Mv class anthocyanins. At the same time, complete shading during the color transition period increases the modification ratio of acylated anthocyanins, enhances the stability of anthocyanin color, and provides a theoretical basis for effectively alleviating the problem of rapid color fading caused by unstable anthocyanin structure in grapes and wines. F3’H and F3’5’H play a crucial role in the biosynthesis pathway of anthocyanins, and the ratio of Cy class anthocyanins catalyzed by F3’H to Dp class anthocyanins catalyzed by F3’5’H affects the color of fruit peels. This study found that during the early stage of color transformation, when shading and harvesting, the expression of F3’H was up-regulated and the expression of F3’5’H was down regulated.During the complete color transition period of shading, both F3’H and F3’5’H were up regulated during harvesting. However, the Cy and Dp anthocyanin contents in the shading treatment during the initial color transition period were significantly higher than those in the shading treatment during the complete color transition period. Perhaps due to the significant positive correlation between CHI expression level Cy and Dp content, and the significantly higher CHI expression level in shading treatment during the early stage of color conversion compared to shading treatment during the complete color conversion period, the Cy and Dp content in shading treatment during the early stage of color conversion was significantly higher than that during shading treatment during the complete color conversion period.There may be differential expression of other key genes, leading to differences in Cy and Dp content, which requires further research.
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