Exploration Of Key Genes And Regulatory Networks Involved In The Development Of Chili Pepper Flowers And Fruits

The development of plant flowers and fruits has always been one of the key issues in breeding production and basic biological research.As an important food and industrial raw material,chili peppers have high economic value.With the rapid development of high-throughput sequence technology,multi-omics techniques have provided a multidimensional global view for the regulatory mechanisms of the development and important quality traits formation of floral organs,seed development,and fruit development in crops such as rice,maize,tomato,strawberry,kiwi,and apple.In this study,we comprehensively analyzed the transcriptome data of 10 stages of chili pepper flower development,as well as the transcriptome and metabolome data of the pericarp,placenta,and seeds at 8stages of development in chili peppers and tomatoes.Through various analysis methods such as WGCNA and K-means clustering,we systematically studied the key genes and important quality trait formation in chili pepper flower and fruit development.The specific research results are as follows:(1)Gene mining and regulatory network research on flower organ morphology construction in pepper.Based on the transcriptome data of 10 developmental stages of chili pepper flower organs and the rich nutritional tissue,at least 30,016 genes were found to be expressed in at least one sample.Using K-means clustering analysis based on the dynamic expression profiles of all genes,the data were divided into 16 clusters with different expression patterns.Through comprehensive differential expression analysis at the whole-genome level,a gene set with specific expression at different stages of chili pepper flower development was identified.Functional enrichment analysis was performed on 333 genes that were specifically upregulated during chili pepper tetramer development.In addition,based on the "ABCDE model" related genes studied in Arabidopsis and tomato,17 candidate genes for the chili pepper "ABCDE model" were identified.They were classified based on their functions in flower organ morphogenesis and a correlation networks map were constructed based on the similarity with their dynamic expression patterns.Six genes were selected to validate the reliability of the transcriptome data.(2)Identification of pepper GTP＿EFTU gene and its function analysis in flower and fruit development.Through analysis at the whole-genome level in chili peppers,a total of 39 genes containing the GTP＿EFTU conserved domain were identified.Combined with the third-generation sequence data of chili pepper tissues at different stages,new information was provided for these genes,including annotations,chromosome positions,phylogenetic relationships,promoter cis-elements,and gene expression patterns in flowers and fruits.GO functional analysis was also performed on co-expressed genes in flower and fruit tissues.(3)Multi-omics association and regulatory network study of capsicum fruit.As a control,the model plant tomato,which has fleshy fruits,was used for comparison.Samples of pericarp,placenta,and seeds were collected from tomato and chili pepper fruits at 8 major developmental stages,and broad-target metabolomics and transcriptomics were performed to construct the full gene expression and metabolite quantification profiles of fruit development in both species.All metabolites were reclassified into13 major categories according to their chemical properties.The distribution patterns of these 13 categories of metabolites in different tissues at 8 stages of fruit development were analyzed,and strong tissue-specific accumulation was observed.K-means clustering analysis and WGCNA were used to perform correlation analysis on the metabolome and transcriptome,and17,303 pairs of direct homologous genes were identified through collinearity analysis in tomato and chili pepper.Based on the dynamic patterns of metabolite and gene expression in the dataset,these metabolites and genes were divided into 12 corresponding clusters.The accumulation of several types of metabolites closely related to fruit quality,such as sugars,organic acids,and alkaloids,were compared and analyzed dynamically,globally demonstrating the similarities and differences in the accumulation of key metabolites in tomato and chili pepper.(4)Mining and verification of key genes in pepper fruit development.Through the analysis of the accumulation pattern of capsaicinoids metabolites and the correlation with the structural genes involved in the capsaicin synthesis pathway,a novel transcription factor Ca MYB15 involved in the regulation of capsaicin synthesis was discovered.It was found that this gene had a strong positive correlation with the known capsaicin synthesis structural genes ACL,4CL,BCKDH,ACS1,KR,and kaslllb.In addition,Ca MYB15 also showed a strong positive correlation with Ca MYB31,a previously validated positive regulator of capsaicin synthesis.VIGS silencing experiments showed that the capsaicinoids content in pepper fruits significantly decreased after the silencing of Ca MYB15,which was found to be localized only in the nucleus.On the other hand,the accumulation of vitamin C in tomato and pepper was compared,and the results showed that both species had the highest vitamin C content in the fruit skin,but overall,the vitamin C content in pepper was about 10 times that of tomato.By homology analysis,candidate genes for vitamin C synthesis pathway in tomato and pepper were identified,and 7differentially expressed genes were found by comparing their expression levels,which provided guidance for further exploration of the differences in vitamin C synthesis between tomato and pepper.