A genetic map of candidate genes and QTLs involved in tomato fruit size and composition

In order to screen for putative candidate genes linked to tomato fruit weight and to sugar or acid content, genes and QTLs involved in fruit size and composition were mapped. Genes were selected among EST clones in the TIGR tomato EST database (http://www.tigr.org/tdb/tgi/lgi/) or corresponded to genes preferentially expressed in the early stages of fruit development. These clones were located on the tomato map using a population of introgression lines (ILs) having one segment of Lycopersicon pennellii (LA716) in a L. esculentum (M82) background. The 75 ILs allowed the genome to be segmented into 107 bins. Sixty-three genes involved in carbon metabolism revealed 79 loci. They represented enzymes involved in the Calvin cycle, glycolysis, the TCA cycle, sugar and starch metabolism, transport, and a few other functions. In addition, seven cell-cycle-specific genes mapped into nine loci. Fourteen genes, primarily expressed during the cell division stage, and 23 genes primarily expressed during the cell expansion stage, revealed 24 and 26 loci, respectively. The fruit weight, sugars, and organic acids content of each IL was measured and several QTLs controlling these traits were mapped. Comparison between map location of QTLs and candidate gene loci indicated a few candidate genes that may influence the variation of sugar or acid contents. Furthermore, the gene/QTL locations could be compared with the loci mapped in other tomato populations.     

Fine mapping of QTLs of chromosome 2 affecting the fruit architecture and composition of tomato

Negative correlations between quality traits and fruit size may hamper the breeding of fresh market tomato varieties for better organoleptic qualities. In a recent QTL analysis, QTLs with large effects on fruit weight, locule number and several quality traits were detected in the distal 50 cM of chromosome 2, but favorable alleles for fruit weight and locule number were unfavorable to quality traits. Substitution mapping was undertaken to determine whether the effects were due to a single QTL or to several tightly linked QTLs. Several chromosomal segments were characterized using near-isogenic lines. Five of them appeared to be involved in one or several traits. Considering the five segments from the top to the bottom of the region, the QTLs detected in each segment controlled the variation of: (1) fruit weight, (2) soluble solids content and dry matter weight, (3) fruit weight, (4) locule number and (5) fruit weight, dry matter weight, total sugars, titratable acidity and soluble solids content. This last cluster illustrates an antagonism between fruit weight and four quality traits, as favorable alleles are not conferred by the same parent in both cases. Nevertheless, several antagonistic QTLs were separated from each other in the first four segments, holding the promise for marker-assisted improvement of fruit quality traits without compromising the fruit size.     

Identification and stability of QTLs for fruit quality traits in apple

Breeding for fruit quality traits is complex due to the polygenic (quantitative) nature of the genetic control of these traits. Therefore, to improve the speed and efficiency of genotype selection, attention in recent years has focused on the identification of quantitative trait loci (QTLs) and molecular markers associated with these QTLs. However, despite the huge potential of molecular markers in breeding programmes, their implementation in practice has been limited by the lack of information on the stability of QTLs across different environments and within different genetic backgrounds. Here, we present the results from a comprehensive analysis of the inheritance of fruit quality traits within a population derived from a cross between the apple cultivars ‘Telamon’ and ‘Braeburn’ over two successive seasons. A total of 74 different QTLs were identified for all the major fruit physiological traits including fruit height, diameter, weight and stiffness, flesh firmness, rate of flesh browning, acidity, the oBrix content and harvest date. Seventeen of these QTLs were ‘major’ QTLs, accounting for over 20% of the observed population variance of the trait. However, only one third (26) of the identified QTLs were stable over both harvest years, and of these year-stable QTLs only one was a major QTL. A direct comparison with published QTL results obtained using other populations (King et al., Theor Appl Genet 102:1227–1235, 2001; Liebhard et al., Plant Mol Biol 52:511–526, 2003) is difficult because the linkage maps do not share a sufficient number of common markers and due to differences in the trait evaluation protocols. Nonetheless, our results suggest that for the six fruit quality traits which were measured in all populations, nine out of a total of 45 QTLs were common or stable across all population × environments combinations. These results are discussed in the framework of the development and application of molecular markers for fruit quality trait improvement.     

Identification and validation of QTLs for salt tolerance during vegetative growth in tomato by selective genotyping.

The purpose of this study was to identify quantitative trait loci (QTLs) for salt tolerance (ST) during vegetative growth (VG) in tomato by distributional extreme analysis and compare them with the QTLs previously identified for this trait. A BC1 population (N = 792) of a cross between a moderately salt-sensitive Lycopersicon esculentum Mill. breeding line (NC84173, maternal and recurrent parent) and a salt-tolerant L. pimpinellifolium (Jusl.) Mill. accession (LA722) was evaluated for ST in solution cultures containing 700 mM NaCl + 70 mM CaCl2 (electrical conductivity, EC = 64 dS/m and phiw approximately -35.2 bars). Thirty-seven BC1 plants (4.7% of the total) that exhibited the highest ST were selected (referred to as the selected population), grown to maturity in greenhouse pots and self-pollinated to produce BC1S1 progeny seeds. The 37 selected BC1S1 progeny families were evaluated for ST and their average performance was compared with that of the parental BC1 population before selection. A realized heritability of 0.50 was obtained for ST in this population. The 37 selected BC1 plants were subjected to restriction fragment length polymorphism (RFLP) analysis using 115 markers, and marker allele frequencies were determined. Allele frequencies for the same markers were also determined in an unselected BC1 population (N = 119) of the same cross. A trait-based marker analysis (TBA), which measures differences in marker allele frequencies between selected and unselected populations, was used to identify marker-linked QTLs. Five genomic regions were detected on chromosomes 1, 3, 5, 6, and 11 bearing significant QTLs for ST. Except for the QTL on chromosome 3, all QTLs had positive alleles contributed from the salt tolerant parent LA722. Of the five QTLs, three (those on chromosomes 1, 3, and 5) were previously identified for this trait in another study, and thus were validated here. Only one of the major QTLs that was identified in our previous study was not detected here. This high level of conformity between the results of the two studies indicates the genuine nature of the identified QTLs and their potential usefulness for ST breeding using marker-assisted selection (MAS). A few BC1S1 families were identified with most or all of the QTLs and with a ST comparable to that of LA722. These families should be useful for the development of salt tolerant tomato lines via MAS.     

Respiration and growth of tomato fruit

The respiration rate and diameter expansion growth of young tomato fruit were measured simultaneously and related to changes in carbon import and plant water status. Respiration rate was directly proportional to the volume expansion rate of fruit growing on isolated plant tops at a positive water potential, whether the growth rate was changed by changing the fruit temperature or by manipulating the source:sink ratio of the plants. From the latter relationship, the maintenance respiration rate was estimated by extrapolation to zero growth and was found to be about 25% of the respiration rate of the average fruit at 21°C. Alternatively, when carbon import was prevented by heat-ringing the fruit peduncle, the respiration rate of the fruit declined to about 40% of the control rate and remained steady, while the expansion rate then declined steadily to >10% of the control rate. These results show that fruit expansion was not contributing significantly to fruit respiration. Indeed, large fluctuations in fruit expansion rate could also be induced by repeated darkening and illumination of potted plants without a corresponding change in fruit respiration. Most significantly, fruit expansion was considerably reduced when plants were allowed to wilt, hut there was no change in fruit respiration rate unless the fruit peduncle was subsequently heat-ringed. We conclude that a major part of the respiration of young tomato fruit was determined by the rate of carbon import, or associated processes, and that fruit expansion per se can occur with relatively low respiratory costs.     

Identification of QTLs for Ralstonia solanacearum race 3-phylotype II resistance in tomato

Resistance against a Ralstonia solanacearum race 3-phylotype II strain JT516 was assessed in a F_2:3 and a population of inbred lines (RIL), both derived from a cross between L. esculentum cv. Hawaii 7996 (partially resistant) and L. pimpinellifolium WVa700 (susceptible). Resistance criteria used were the percentage of wilted plants to calculate the AUDPC value, and bacterial colonization scores in roots and stem (hypocotyl and epicotyl) assessed in two independent greenhouse experiments conducted during the cool and hot seasons in Réunion Island, France. Symptoms were more severe during the cool season trials. Heritability estimates in individual seasons ranged from 0.82 to 0.88, depending on resistance criterion. A set of 76 molecular markers was used for quantitative trait loci (QTL) mapping using the single- and composite- interval mapping methods, as well as ANOVA. Four QTLs, named Bwr- followed by a number indicating their map location, were identified. They explained from 3.2 to 29.8% of the phenotypic variation, depending on the resistance criterion and the season. A major QTL, Bwr-6, and a minor one, Bwr-3, were detected in each season for all resistance criteria. Both QTLs showed stronger effects in the hot season than in the cool one. Their role in resistance to R. solanacearum race 3-phylotype II was subsequently confirmed in the RIL population derived from the same cross. Two other QTLs, Bwr-4 and Bwr-8, with intermediate and minor effects, respectively, were only detected in the hot season, demonstrating that environmental factors may strongly influence the expression of resistance against the race 3-phylotype II strain JT516. These QTLs were compared with those detected in the RIL population against race 1-phylotype I strain JT519 as well as those detected in other previous studies in the same genetic background against other race 1-phylotype I and II strains. This comparison revealed the possible occurrence of some phylotype-specific resistance QTLs in Hawaii 7996.     

ABA-deficiency results in reduced plant and fruit size in tomato

Abscisic acid (ABA) deficient mutants, such as notabilis and flacca, have helped elucidating the role of ABA during plant development and stress responses in tomato (Solanum lycopersicum L.). However, these mutants have only moderately decreased ABA levels. Here we report on plant and fruit development in the more strongly ABA-deficient notabilis/flacca (not/flc) double mutant. We observed that plant growth, leaf-surface area, drought-induced wilting and ABA-related gene expression in the different genotypes were strongly correlated with the ABA levels and thus most strongly affected in the not/flc double mutants. These mutants also had reduced fruit size that was caused by an overall smaller cell size. Lower ABA levels in fruits did not correlate with changes in auxin levels, but were accompanied by higher ethylene evolution rates. This suggests that in a wild-type background ABA stimulates cell enlargement during tomato fruit growth via a negative effect on ethylene synthesis.     

A histidine decarboxylase-like mRNA is involved in tomato fruit ripening

DNA sequencing of a tomato ripening-related cDNA, TOM 92, revealed an open reading frame with homology to several pyridoxal 5-phosphate histidine decarboxylases, containing the conserved amino acid residues known to bind pyridoxal phosphate and -fluoromethylhistidine, an inhibitor of enzyme activity. TOM 92 mRNA accumulated during early fruit ripening and then declined. Fruit of the ripeningimpaired tomato mutant, ripening inhibitor (rin), did not accumulate TOM 92 mRNA, and its accumulation was not restored by treatment of fruit with ethylene. The TOM 92 mRNA was not detected in tomato leaves and unripe fruit.     

Molecular expression of genes involved in parthenocarpic fruit set in tomato

To characterize the phenomenon of natural parthenocarpy in tomato ( Lycopersicon esculentum Mill.) two different approaches have been followed. At a developmental level, the ovary weights of three non-parthenocarpic lines and three near-isogenic parthenocarpic ( pat-2 ) lines were compared. Four developmental stages were considered: flower bud, preanthesis, anthesis and 4 days after anthesis. The parthenocarpic lines displayed ovary weights higher than their respective non-parthenocarpic lines from preanthesis to 4 days after anthesis. A molecular approach involved comparison of in vitro translation products from flower RNAs taken from the same developmental stages of non-parthenocarpic and near-isogenic parthenocarpic ( pat-2 and pat-3/pat-4 ) lines. Analysis by two-dimensional polyacrylamide gel electrophoresis showed the differential expression of a 30-kDa product in parthenocarpic materials from preanthesis to anthesis. These results suggest that the physiological and molecular events responsible for parthenocarpy begin at the preanthesis stage, before the flower is completely mature and receptive to pollination. The differential expression of this in vitro translation product in pat-2 and pat-3/pat-4 genotypes also suggests a common or confluent molecular basis in genetically controlled parthenocarpy.     

Relationship between tomato fruit growth and fruit osmotic potential under salinity

To investigate the relationship between fruit growth and fruit osmotic potential (Ψ_s) in salty conditions, a sensitive tomato cultivar (Lycopersicon esculentum Mill.) and a tolerant accession of the wild species Lycopersicon pimpinellifolium Mill. were grown in a greenhouse with 0 and 70 mM NaCl, and the growth of the fruit studied from 15 to 70 days after anthesis (DAA). L. pimpinellifolium did not reduce significantly fruit weight in salty conditions throughout the growth period, whereas L. esculentum fruit weights decreased significantly with salinity from 45 DAA. L. esculentum fruit fresh weight reductions resulted from both less dry matter and water accumulation, although the fruit water content was affected by salinity before the fruit weight. In both species, fruit osmotic potential (Ψ_s) decreased significantly with salinity during the rapid fruit growth phase, although the changes were different. Thus, fruits from L. pimpinellifolium salt treated plants showed a Ψ_s reduction at the beginning (15 DAA) twice as high as that found in L. esculentum. As the advanced growth stage (from 15 to 55 DAA), the Ψ_s reduction percentages induced by salinity were quite similar in L. pimpinellifolium fruits, while increased in L. esculentum. Under saline conditions, the solutes contributing to reduce the fruit Ψ_s during the first 55 DAA were the inorganic solutes in both species, while in the ripe fruits they were hexoses. L. esculentum fruits accumulated K⁺ as the main osmoticum in salty conditions, while L. pimpinellifolium fruits were able to use not only K⁺ but also the Na⁺ provided by the salt.     

Marker-assisted introgression of five QTLs controlling fruit quality traits into three tomato lines revealed interactions between QTLs and genetic backgrounds

The evaluation of organoleptic quality of tomato fruit requires physical, chemical and sensory analyses, which are expensive and difficult to assess. Therefore, their practical use in phenotypic selection is difficult. In a previous study, the genetic control of several traits related to organoleptic quality of fresh-market tomato fruit was investigated. Five chromosome regions strongly involved in organoleptic quality attributes were then chosen to be introgressed into three different recipient lines through marker-assisted selection. A marker-assisted backcross (MABC) strategy was performed, as all the favorable alleles for quality traits were provided by the same parental tomato line, whose fruit weight (FW) and firmness were much lower than those of the lines commonly used to develop fresh market varieties. Three improved lines were obtained after three backcrossing and two selfing generations. The implementation of the MABC scheme is described. The three improved lines were crossed together and with the recipient lines in a half-diallel mating scheme, and the simultaneous effect of the five quantitative trait locus (QTL) regions was compared in different genetic backgrounds. Significant effects of the introgressed regions and of the genetic backgrounds were shown. Additive effects were detected for soluble solid and reducing sugar content in two genetic backgrounds. A partially dominant effect on titratable acidity was detected in only one genetic background. In contrast, additive to dominant unfavorable effects of the donor alleles were detected for FW and locule number in the three genetic backgrounds. Recessive QTL effects on firmness were only detected in the two firmest genetic backgrounds. Comparison of the hybrids in the half-diallel gave complementary information on the effects of: (1) the alleles at the selected regions, (2) the genetic backgrounds and (3) their interaction. Breeding efficiency strongly varied according to the recipient parent, and significant interactions between QTLs and genetic backgrounds were shown for all of the traits studied.     

Identifying the loci responsible for natural variation in fruit size and shape in tomato

Fruit size and shape are two major factors determining yield, quality and consumer acceptability for many crops. Like most traits important to agriculture, both are quantitatively inherited. Despite their economic importance none of the genes controlling either of these traits have been cloned, and little is known about the control of the size and shape of domesticated fruit. Tomato represents a model fruit-bearing domesticated species characterized by a wide morphological diversity of fruits. The many genetic and genomic tools available for this crop can be used to unraveal the molecular bases of the developmental stages which presumably influence fruit architecture, size and shape. The goal of this review is to summarize data from the tomato QTL studies conducted over the past 15 years, which together allow the identification of the major QTLs responsible for fruit domestication in tomato. These results provide the starting point for the isolation of the genes involved in fruit-size/shape determination in tomato and potentially other fruit-bearing plants. Received: 21 January 1999 / Accepted: 12 March 1999     

Comparison of ripening processes in intact tomato fruit and excised pericarp discs

Physiological processes characteristic of ripening in tissues of intact tomato fruit (Lycopersicon esculentum Mill.) were examined in excised pericarp discs. Pericarp discs were prepared from mature-green tomato fruit and stored in 24-well culture plates, in which individual discs could be monitored for color change, ethylene biosynthesis, and respiration, and selected for cell wall analysis. Within the context of these preparation and handling procedures, most whole fruit ripening processes were maintained in pericarp discs. Pericarp discs and matched intact fruit passed through the same skin color stages at similar rates, as expressed in the L^*a^*b^* color space, changing from green (a^* < −5) to red (a^* > 15) in about 6 days. Individual tissues of the pericarp discs changed color in the same sequence seen in intact fruit (exocarp, endocarp, then vascular parenchyma). Discs from different areas changed in the same spatial sequence seen in intact fruit (bottom, middle, top). Pericarp discs exhibited climacteric increases in ethylene biosynthesis and CO₂ production comparable with those seen in intact fruit, but these were more tightly linked to rate of color change, reaching a peak around a^* = 5. Tomato pericarp discs decreased in firmness as color changed. Cell wall carbohydrate composition changed with color as in intact fruit: the quantity of water-soluble pectin eluted from the starch-free alcohol insoluble substances steadily increased and more tightly bound, water-insoluble, pectin decreased in inverse relationship. The cell wall content of the neutral sugars arabinose, rhamnose, and galactose steadily decreased as color changed. The extractable activity of specific cell wall hydrolases changed as in intact fruit: polygalacturonase activity, not detectable in green discs (a^* = −5), appeared as discs turned yellow-red (a^* = 5), and increased another eight-fold as discs became full red (a^* value +20). Carboxymethyl-cellulase activity, low in extracts from green discs, increased about six-fold as discs changed from yellow (a^* = 0) to red.     

Antisense acid invertase (TIV1) gene alters soluble sugar composition and size in transgenic tomato fruit.

Invertase (beta-fructosidase, EC 3.2.1.26) hydrolyzes sucrose to hexose sugars and thus plays a fundamental role in the energy requirements for plant growth and maintenance. Transgenic plants with altered extracellular acid invertase have highly disturbed growth habits. We investigated the role of intracellular soluble acid invertase in plant and fruit development. Transgenic tomato (Lycopersicon esculentum Mill.) plants expressing a constitutive antisense invertase transgene grew identically to wild-type plants. Several lines of transgenic fruit expressing a constitutive antisense invertase gene had increased sucrose and decreased hexose sugar concentrations. Each transgenic line with fruit that had increased sucrose concentrations also had greatly reduced levels of acid invertase in ripe fruit. Sucrose-accumulating fruit were approximately 30% smaller than control fruit, and this differential growth correlated with high rates of sugar accumulation during the last stage of development. These data suggest that soluble acid invertase controls sugar composition in tomato fruit and that this change in composition contributes to alterations in fruit size. In addition, sucrose-accumulating fruit have elevated rates of ethylene evolution relative to control fruit, perhaps as a result of the smaller fruit size of the sucrose-accumulating transgenic lines.     

Water relations and growth of tomato fruit pericarp tissue

The water relations of young tomato fruit pericarp tissue were examined and related to tissue expansion. The relationship between bulk turgor pressure and tissue expansion (as change in fresh mass or length of tissue) was determined in slices of pericarp cut from young, growing fruit by incubation in different osmotic concentrations of polyethylene glycol 6000 or mannitol. The bulk turgor of this tissue was low (about 0.2 MPa), even in fruit from plants that were otherwise fully turgid, whether measured psychrometrically or by length change in osmotic solutions. The rate of tissue growth at maximum turgor was less than that at moderate turgor unless calcium was added to the incubation medium. However, added calcium also decreased the rate of growth at lower turgor pressures. Yield turgor was < 0.1 MPa, but it was increased by the addition of calcium ions. Electrolyte leakage from tissue was greatest at maximum turgor pressure but was decreased by the addition of calcium ions or osmoticum. Tissue growth was unaffected by a range of plant growth regulators (IAA, abscisic acid, benzyladenine and GA₃) but was inhibited, particularly at high turgor, by low concentrations of malic or citric acid. The low turgor pressure of pericarp tissue could be due to the presence of apoplastic solutes within the pericarp, and evidence for this is discussed. Thus, fruit tissue may be able to maintain optimal expansion rates only at moderate turgor and low calcium concentration.     

Water relations of the tomato during fruit growth

Fruit and stem water potentials of tomato plants were measured continuously for several days using automated psychrometers. A linear voltage displacement transducer was used to simultaneously measure diameter changes on an adjacent fruit. A strong correlation was observed between the water potential gradient of the fruit and stem, and changes in fruit diameter. Fruit diameter increased when the apoplasmic water potential gradient favoured solution flow into the fruit and fruit shrinkage occurred only when the water potential gradient was inverted. Based on our data and other published data (Ehret & Ho 1986; Lee 1989a) on phloem transport in tomato, we have concluded that low stem water potentials have an immediate and direct effect on phloem turgor; reducing the driving force for sap flow into the fruit. Since fruit water potential remained relatively constant, the diurnal variation in stem water potential was sufficient to account for the correlation with changes in fruit diameter. There are consequences with respect to predicting the accumulation of dry matter in tomato fruit.