Transcriptome Analysis Of Different Fruit-shaped Grape(Vitis Vinifera L.) And Function Analysis Of TIFY10A Gene

Grapes（Vitis vinifera）belong to the grape family,woody vines deciduous plants.In our country,the grape industry is mainly table grape variety,its fruit character determines the quality and commodity value.The shape of grape fruit also directly affects its economic value because of different consumption habits.Although the existing grape varieties are rich in fruit shape,it is rare to study the internal mechanism of fruit shape difference.In this study,three kinds of table grape varieties with large differences in fruit shape:oblong variety?Rizamat’,ovate variety?Victoria’and suborbicular variety?Fujiminori’were used as experimental materials.By observing the different fruit-shape of cell morphology and determine endogenous hormone content,using the transcriptome sequencing analysis,screening fruit-shape formation regulatory genes for cloning and transformation,the preliminary analysis to its function,from the physiological and molecular level,aims to explore the internal mechanism of grape fruit-shape formation from the physiological and molecular aspects.The main findings are as follows:1.The phenotypic analysis of the young fruit stage,expansion stage and veraison of different fruit shape grapes found that the longitudinal and horizontal diameters of the three grape varieties showed a significant increase trend,and all reached the highest in the veraison.At the young fruit stage,there were no significant differences in the longitudinal and horizontal diameters of the three grape varieties,but the oblong variety?Rizamat?had higher levels of Zeatin（ZT）and Gibberellin acid（GA₃）in the fruits.At the expansion period,the longitudinal diameter order is?Victoria?>?Rizamat?>?Fujiminori’,the horizontal diameter order is?Victoria?>?Fujiminori’>?Rizamat?,and the difference is significant while ZT and GA₃ decreased most significantly in the fruit of?Rizamat?.At the veraison,the longitudinal diameter order is?Rizamat?>?Victoria?>?Fujiminori’,the horizontal diameter order is?Fujiminori’>?Victoria?>?Rizamat?,and the difference is significant,and the content of IAA,GA₃ and ZT in the fruits of?Fujiminori’was the highest during this period.In the veraison,the fruit shape index is in the order of?Rizamat?>?Victoria?>?Fujiminori’≈1,and the fruit shape of?Fujiminori’is more similar to a circle.Observation of the cell morphology of the pulp microstructure revealed that there are significant differences in the cell length and per unit area cell number of the three varieties of grapes in the expansion stage,among them,the length of pulp cells order is‘Rizamat’>’Victoria’>’Fujiminori’,the number of per unit area cells order is:‘Victoria’>’Rizamat’>’Fujiminori’,and there are significant differences.For the oblong variety‘Rizamat’and the ovate variety‘Victoria’,compared to the suborbicular variety‘Fujiminori’,the pulp cells tend to grow more elongated,and the cells of‘Victoria’and?Fujiminori?are more densely arranged,to sum up,we preliminarily speculate that in different fruit-shaped grapes,cell morphology,cell number and endogenous hormone levels have a potential impact on the formation of fruit shape.2.Perform transcriptome sequencing（RNA-Seq）for different fruit-shaped grape varieties before the fruit morphology is established（young fruit stage）,a total of 2288differentially expressed genes（DEGs）were identified.There were 1285 DEGs in the oblong vs suborbicular sample,673 were up-regulated and 612 were down-regulated;909 DEGs in the ovate vs oblong sample,424 were up-regulated and 482 were down-regulated;1302 DEGs in the ovate vs suborbicular sample,620 were up-regulated and682 were down-regulated.It showed that the number of DEGs in near-round and non-round grapes is slightly higher.KEGG was significantly enriched in the"plant hormone signal transduction"pathway,and 20 DEGs were identified,based on their expression patterns and transcription factor analysis,the Vv TIFY10A gene was screened.The expression level of this gene in the oblong variety‘Rizamat’was significantly up-regulated compared to the ovate variety‘Victoria’,while the expression level of this gene in the suborbicular‘Fujiminori’was significantly lower than that of the ovate variety‘Victoria’;it is speculated that grape fruit formation may be related to DEGs in the"plant hormone signal transduction"pathway.And using real-time fluorescence quantitative（q RT-PCR）method to verify the transcriptome data,the results show that the data is reliable.Analyze the function of Vv TIFY10A gene by constructing Vv TIFY10A-GFP over-expression vector.By subcellular localization of onion epidermis and tobacco leaves,the results showed that the Vv TIFY10A gene was localized in the nucleus,which was consistent with the results of bioinformatics prediction.It was further transformed into tomato（Micro-Tom）,in over-expressed Vv TIFY10A tomato,the longitudinal and horizontal diameter of the fruit increased by 8.17%and 16.37%compared with the WT.Preliminary speculation that this gene may regulate the shape of the fruit.