The Research On The Effect Of Calcium On The Development Of Hazel Fruits And The Calcium-Related Gene Regulatory Network

Fruit abortion is one of the main factors affecting the hazelnut yields in China,however,the genetic mechanism of hazelnut abortion is not clear.Some reports show that calcuim has important effect on the growth of other plants.To explore the physiological and molecular mechanism of calcium in the development of hazelnut fruit,this experiment which was based on the local main variety as the test material,was made to analyze the molecular mechanism leading to fruit abortion and the calcium-related genes regulating fruit development by comparing the differences between transcriptome and proteome in normal and abortive fruit.At the same time,this experiment used a weighted gene co-expression analysis method to construct a calcium regulation network at different stages,and described the physiological activities of calcium involved in various periods to provide a theoretical basis for the study of the mechanism of calcium on fruit development.In addition,the effects of calcium treatment on calcium regulation network were further verified by gene expression profiles under different calcium treatment conditions,and the effects of different concentrations of CaCl2 on fruit growth and yield were verified.Some conclusions of bioinformatics were verified simultaneously.(1)The molecular mechanism responsible for fruit abortion was obtained by transcriptome and proteomic analysis.In this experiment,the normal fruit was used as the control,the aborted fruit was treated,and the transcriptome and protein group were sequenced.In the transcriptome sequencing,2970 up-regulated genes and 1384 down-regulated genes were obtained.GO and KEGG enrichment were performed on up-regulated and down-regulated differentially expressed genes respectively.The results showed that the chloroplast and photosynthesis genes were significantly expressed in normal fruits.In proteomic analysis,237 differentially expressed proteins were up-regulated in abortive fruits,and 376 differentially expressed proteins were down-regulated.GO and KEGG enrichment analysis of differentially expressed proteins up-regulated and down-regulated were performed.The results showed that the most of the differentially expressed proteins enriched in normal fruits were associated with photosynthesis,hormone-related pathways and stress response.A correlation analysis between differentially expressed genes and differentially expressed proteins was performed.8 GO term and 3 KEGG pathways in which genes/proteins highly expressed in normal fruits were significantly enriched in RNA and protein levels.(2)Calcium-related genes that play important regulatory roles during fruit development were obtained.Differentially expressed EF-hand calcium-binding protein gene was obtained by transcriptome analysis.Calmodulin,calmodulin-like,and calcium/calmodulin-dependent protein kinases were up-regulated in abortive fruits.In the proteome analysis,31 calcium-related differentially expressed proteins that may be involved in the development of hazelnut,including calmodulin,calmodulin-like,calcium/calmodulin-dependent protein kinase,etc.,were found.Three calcium-related genes were simultaneously highly expressed at mRNA/protein levels,including calmodulin,calcium-sensitive receptor and Photosystem II aerator enhancer protein 2 by correlation analysis of calcium-related proteins in the transcriptome and proteome.Associated genes/proteins played an important role in fruit development.(3)The calcium regulation network of different stages was constructed,and the physiological activities of calcium in various periods were described.The expression analysis was used to further verify the effect of calcium treatment on calcium regulation network.Using transcriptome data from unpollinated pistil sample(F),primary pistil inflorescence sample(S),pistil inflorescence sample formed early ovule(T),fruit cluster samples completed ovule(F0)performed WGCNA analysis.The results showed:among the 22 modules,the brown module was closely related to the T,and the yellow module was closely related to the S.The first three modules of calcium signal transduction-related gene enrichment were blue module,red module and brown module by analysis of the distribution of calcium-related genes in each module.GO and KEGG enrichment analysis were performed on genes enriched in different modules,and the functions and metabolic pathways corresponding to each module gene were interpreted to understand the calcium-related signal regulation network and its role in the development of fruit.The pollinated hazelnuts were sprayed with different concentrations of CaCl2,and yellow module of the S was selected for verification of the regulatory network.The results show the credibility of the regulatory network.(4)Spraying calcium can significantly increase the calcium content and the fruit diameter of the young fruits.At the flowering stage,spraying calcium can increase the average fruit number and fruit setting rate.There was a significant positive correlation between the calcium content of young fruit and the average fruit number.At the same time,there was a significant positive correlation between the calcium content of young fruit and fruit setting rate.There was a significant negative correlation between the calcium content of young fruit and the number and abortion rate of abortive fruit.Partial conclusions from bioinformatics were verified.