Breeding Tomato for Increased Fruit Phenolics

The purpose of this research was to develop tools and germplasm for the production of tomatoes (Solanum lycopersicum L.) with high fruit phenolics without the use of genetic engineering. A candidate gene analysis was undertaken to identify the genes Anthocyanin fruit tomato (Aft) and Purple Smudge, which regulate anthocyanin biosynthesis in tomato fruit. Prevously, we demonstrated that Aft co-segregated with a DNA polymorphism in a Myb transcription factor (SlAn2) similar to Petunia An2. Expression analysis of the SlAn2 gene using semi-quantitative reverse transcription PCR showed a close correlation between transcript levels of SlAn2, the anthocyanin biosynthetic gene dihydroflavonol 4-reductase (DFR), and anthocyanin expression. Expression of anthocyanin in fruit of Aft and Purple Smudge genotypes was associated with increased expression of SlAn2. Expression analysis of the related Myb transcription factor SlAnt1 showed poor correlation with anthocyanin expression in the fruit, indicating that SlAn2 is a better candidate gene for the Aft and Purple Smudge genes than SlAnt1. Phylogenetic analysis of the predicted amino acid sequences of tomato SlAnt1 and SlAn2 indicated that they are orthologous to potato StAn1 and StAn2, respectively. To create a high flavonol tomato line, an elite high anthocyanin line with the genes Aft and atv was crossed to a tomato line with the gene anthocyanin without (aw), which encodes a non-functional DFR. Aft/atv/aw F₃ segregants had fruit with levels of rutin and chlorogenic acid that were significantly higher than in the cultivar ‘Legend’ under greenhouse conditions. Total phenolics levels in fruit of Aft/atv/aw F₃ segregants were not significantly different from an elite Aftatv line under field conditions. However, small amounts of anthocyanin were unexpectedly found in Aft/atv/aw fruit.  In order to identify and introgress additional genes that would increase total phenolics or result in novel phenolic compounds in tomato fruit, we screened a core collection of Solanum lycopersicum var. cerasiforme Dunal. Several S.l. var. cerasiforme accessions with good horticultural quality and significantly higher levels of total phenolics in fruit were identified. Some of the S.l. var. cerasiforme accessions also contained novel polyphenolic compounds not detected in cultivated tomato fruit. These S.l. var. cerasiforme accessions have been crossed to elite high anthocyanin and high flavonol tomato lines and will be extensively genotyped by the SolCAP project, facilitating the introgression of genes from them.