Dissecting Tomato Fruit Resistance Andsoluble Solid Content Genes By Constructing Pan-transcriptome

Tomato is not only a high-value crop worldwide but also a model plant for fleshy fruit development.Tomato stress resistance and fruit total soluble solids(TSS)content have changed dramatically during selective breeding.Here,we constructed a pan-transcriptome of tomato orange-stage fruit,investigated global expression presence/absence variation(ePAV),differential gene expression,and revealed 19 key genes associated with TSS content based on RNA-seq data of 399 tomato accessions.The main results of this study are summarized as follows:1.To profile and quantify RNA transcripts in the whole transcriptome of orange-stage tomato fruit,we performed a population-level transcript assembly based on the ‘Heinz 1706' reference genome and the de novo assembly of genes that were absent from the ‘Heinz 1706' reference genome by RNA-seq of orange-stage fruit from 26 PIM,114 CER,and 259 BIG accessions.Firstly,we performed a population-level transcript assembly.We identified 22,258,24,052,and 25,134 known genes and 2,282,4,730,and 6,657 novel genes expressed in PIM,CER,and BIG,respectively.Genes identified in each group were merged to obtain a population-level transcriptome.In total,34,084 genes were assembled based on the reference genome.Next,reads that failed to map to the ‘Heinz 1706' reference genome sequence were used to perform a de novo assembly of genes that were absent from the reference genome in each group.We finally identified 907,2,328,and 3,946 high-confidence RTAs absent from the ‘Heinz 1706' reference sequence in PIM,CER,and BIG,respectively.Totally,we identified 41,265 genes expressed in tomato fruit at the orange-fruit stage,of which 15411 genes were newly assembled genes.2.To identify the ePAV genes during domestication and breeding,we constructed orthologous gene clusters using OrthoMCL clustering.We identified 6,123 RTAs orthologous clusters,including 2,706 ePAV genes during tomato domestication and 4,992 ePAV genes during tomato breeding.Further,we found that the gene PIM_DN29746_c0_g3_i1 annotated as a late blight resistance protein and the gene PIM_DN30274_c0_g2_i1 encoding peroxidase P7 lost their expression during domestication.3.To dissect the transcriptome-level variation in tomato based on ‘Heinz 1706' reference gene models,we identified differentially expressed genes(DEGs)among the PIM,CER,and BIG groups.The expression level of 3,629 genes was significantly up/down-regulated following domestication or breeding.Among these DEGs,1,945 were differentially expressed upon domestication,and 762 DEGs associated with breeding.Furthermore,we detected 300 genes that showed differential expression throughout domestication and breeding.4.We identified 19 genes were associated with the fruit TSS content,including LIN5,TIV1,and seven novel sugar transporter genes,three of which coding bidirectional sugar transporter genes are located at the quantitative trait locus(QTL)associated with glucose content.In addition,the gene encoding citrate synthase was significantly downregulated upon domestication and breeding.This gene might account for the reduce of fruit TSS content during domestication and breeding.