Genomic Dissection of a Wild Region in a Superior Solanum pennellii Introgression Sub-Line with High Ascorbic Acid Accumulation in Tomato Fruit

Abstract

The Solanum pennellii introgression lines (ILs) have been exploited to map quantitative trait loci (QTLs) and identify favorable alleles that could improve fruit quality traits in tomato varieties. Over the past few years, ILs exhibiting increased content of ascorbic acid in the fruit have been selected, among which the sub-line R182. The aims of this work were to identify the genes of the wild donor S. pennellii harbored by the sub-line and to detect genes controlling ascorbic acid accumulation by using genomics tools. A Genotyping-By-Sequencing (GBS) approach confirmed that no wild introgressions were present in the sub-line besides one region on chromosome 7. By using a dense single nucleotide polymorphism (SNP) map obtained by RNA sequencing (RNA-Seq), the wild region of the sub-line was finely identified; thus, defining 39 wild genes that replaced 33 genes of the ILs genetic background (cv. M82). The differentially expressed genes mapping in the region and the variants detected among the cultivated and the wild alleles evidenced the potential role of the novel genes present in the wild region. Interestingly, one upregulated gene, annotated as a major facilitator superfamily protein, showed a novel structure in R182, with respect to the parental lines. These genes will be further investigated using gene editing strategies.

Keywords: RNA-Seq; Solanum lycopersicum; genome reconstruction; genotyping-by-sequencing; introgression lines; red ripe fruit.

Figure 1

Volcano plots of genes differentially expressed between R182 and M82 at (A) breaker and (B) mature red fruit stages. The y-axis corresponds to the mean expression value of log 10 (false discovery rate, FDR) and the x-axis displays the log2 (fold change) value. The red dots represent the significantly upregulated genes while the green dots represent the significantly downregulated genes. Grey dots represent the genes whose expression levels did not reach statistical significance.

Figure 2

Reconstruction of the introgressed wild region in R182 by the synteny analysis of the genomic portion of chromosome 7 between S. pennellii and S. lycopersicum. The introgressed genes of S. pennellii, the replaced genes of S. lycopersicum, and their function annotation were highlighted in the right side of the figure. Genes evidenced in red represent species-specific genes. Experimentally tested markers (+) and duplicated genes (*) were reported next to the corresponding genes.

Figure 3

Multi-alignment of the last exon of Sopen07g024640, the ortholog Solyc07g049310 and the recombined sequence in R182. The recombination event occurred between the green rectangle showing the last S. pennellii variant of the R182 gene and the red rectangles showing the S. lycopersicum variants of R182 gene. The blue arrow points out the position where the 3’ Untranslated Region (UTR) starts.