Study On Fruit Development And Quality Formation Of Pumpkin Based On Transcriptome

Pumpkin is one of the main vegetable crops of Cucurbita,which is widely cultivated in the world.China’s pumpkin cultivation area and output are in the world’s leading position,and pumpkin industry has become one of the industry pillars for farmers to get rich.With the continuous expansion of cultivation area and industrial field,the requirements for pumpkin quality are constantly improved.Identification of candidate genes related to fruit development and quality formation is an effective way to improve the quality of pumpkin fruit.In this study,therefore,the pulp of Cucurbita maxima Duch.inbred lines(“Rimu”)were collected at different development period(0,10,20,30,40 and 50 days after pollination).First,fruit nutrients at different stages were determined.Then RNA-Seq was used to systematically identify the differentially expressed genes related to fruit development and quality.This study provides an effective basis for pumpkin quality precision breeding and the mining of important candidate genes,and also lays a theoretical foundation for the study of the regulation mechanism of pumpkin quality metabolism.The main results are as follows:1.Determination of fruit nutrients in different stages of pumpkin developmentThe content of total protein,starch and soluble sugar in pumpkin fruits after pollination were determined.The results showed that the nutrient content of pumpkin fruits changed with the fruit development in 6 different stages.The change trends of total protein,starch and soluble sugar in pumpkin fruits in different developmental stages were preliminarily obtained.The change of total protein content was first decreased and then increased,while the change law of starch content was“slow,fast,slow and fast ”.The change law of soluble sugar made the continuous of ruit after pollination reach the peak and then decrease.2.Transcriptome sequencing of pumkin fruits and analysis of differentially expressed genes related to fruit developmentUsing RNA-seq technology,high-throughput transcriptome sequencing of pumpkin fruits at different development stages was carried out.After the quality control of sequencing,a total of 298.09 Gb Clean Data was obtained from 18 samples.The Clean Data of each sample reached 16.06 Gb,and the percentage of Q30 base was93.57% or above.Sequence alignment of Clern Reads from each samples with reference genome ranged from 88.92% to 95.52%.Based on the comparison results,geneexpression analysis was conducted.According to the gene expression in differnet samples,15,307 differentially expressed genes were identified,and functional annotation and enrichment analysis were carried out.Among them,944 genes were differentially expressed in 10 or more comparison groups.Combined with gene functional annotation,GO and KEGG analysis,47 significantly differentially expressed enzyme genes related to sugar and starch anabolism were selected as candidate genes.3.Real-time quantitative PCR validation and co-expression analysis of candidaegenesThe expression patterns of the 47 candidate genes were verified by real-timequantitative PCR.The results showed that the expression trend of quantitative PCR results was roughly the same as that of sequencing results,which verified thereliability of transcriptome sequencing data.Co-expression analysis was conductedusing the expression data of the 47 candidate genes,and a total of 49 pairs of genes were found to have interactions,providing indirect evidence for the interactions between the genes in the starch and sugar metabolism pathways.