Dissecting genetic architecture of grape proanthocyanidin composition through quantitative trait locus mapping.
ABSTRACT: BACKGROUND: Proanthocyanidins (PAs), or condensed tannins, are flavonoid polymers, widespread throughout the plant kingdom, which provide protection against herbivores while conferring organoleptic and nutritive values to plant-derived foods, such as wine. However, the genetic basis of qualitative and quantitative PA composition variation is still poorly understood. To elucidate the genetic architecture of the complex grape PA composition, we first carried out quantitative trait locus (QTL) analysis on a 191-individual pseudo-F1 progeny. Three categories of PA variables were assessed: total content, percentages of constitutive subunits and composite ratio variables. For nine functional candidate genes, among which eight co-located with QTLs, we performed association analyses using a diversity panel of 141 grapevine cultivars in order to identify causal SNPs. RESULTS: Multiple QTL analysis revealed a total of 103 and 43 QTLs, respectively for seed and skin PA variables. Loci were mainly of additive effect while some loci were primarily of dominant effect. Results also showed a large involvement of pairwise epistatic interactions in shaping PA composition. QTLs for PA variables in skin and seeds differed in number, position, involvement of epistatic interaction and allelic effect, thus revealing different genetic determinisms for grape PA composition in seeds and skin. Association results were consistent with QTL analyses in most cases: four out of nine tested candidate genes (VvLAR1, VvMYBPA2, VvCHI1, VvMYBPA1) showed at least one significant association with PA variables, especially VvLAR1 revealed as of great interest for further functional investigation. Some SNP-phenotype associations were observed only in the diversity panel. CONCLUSIONS: This study presents the first QTL analysis on grape berry PA composition with a comparison between skin and seeds, together with an association study. Our results suggest a complex genetic control for PA traits and different genetic architectures for grape PA composition between berry skin and seeds. This work also uncovers novel genomic regions for further investigation in order to increase our knowledge of the genetic basis of PA composition.
Project description:Grape proanthocyanidins (PAs) play a major role in the organoleptic properties of wine. They are accumulated mainly in grape skin and seeds during the early stages of berry development. Despite the recent progress in the identification of genes involved in PA biosynthesis, the mechanisms involved in subunit condensation, galloylation, or fine regulation of the spatio-temporal composition of grape berries in PAs are still not elucidated. Two Myb transcription factors, VvMybPA1 and VvMybPA2, controlling the PA pathway have recently been identified and ectopically over-expressed in an homologous system. In addition to already known PA genes, three genes coding for glucosyltransferases were significantly differentially expressed between hairy roots over-expressing VvMybPA1 or VvMybPA2 and control lines. The involvement of these genes in PA biosynthesis metabolism is unclear. The three glucosyltransferases display high sequence similarities with other plant glucosyltransferases able to catalyse the formation of glucose esters, which are important intermediate actors for the synthesis of different phenolic compounds. Studies of the in vitro properties of these three enzymes (K(m), V(max), substrate specificity, pH sensitivity) were performed through production of recombinant proteins in E. coli and demonstrated that they are able to catalyse the formation of 1-O-acyl-Glc esters of phenolic acids but are not active on flavonoids and stilbenes. The transcripts are expressed in the early stages of grape berry development, mainly in the berry skins and seeds. The results presented here suggest that these enzymes could be involved in vivo in PA galloylation or in the synthesis of hydroxycinnamic esters.
Project description:BACKGROUND:High temperature during grape berry ripening impairs the quality of fruits and wines. Veraison time, which marks ripening onset, is a key factor for determining climatic conditions during berry ripening. Understanding its genetic control is crucial to successfully breed varieties more adapted to a changing climate. Quantitative trait loci (QTL) studies attempting to elucidate the genetic determinism of developmental stages in grapevine have identified wide genomic regions. Broad scale transcriptomic studies, by identifying sets of genes modulated during berry development and ripening, also highlighted a huge number of putative candidates. RESULTS:With the final aim of providing an overview about available information on the genetic control of grapevine veraison time, and prioritizing candidates, we applied a meta-QTL analysis for grapevine phenology-related traits and checked for co-localization of transcriptomic candidates. A consensus genetic map including 3130 markers anchored to the grapevine genome assembly was compiled starting from 39 genetic maps. Two thousand ninety-three QTLs from 47 QTL studies were projected onto the consensus map, providing a comprehensive overview about distribution of available QTLs and revealing extensive co-localization especially across phenology related traits. From 141 phenology related QTLs we generated 4 veraison meta-QTLs located on linkage group (LG) 1 and 2, and 13 additional meta-QTLs connected to the veraison time genetic control, among which the most relevant were located on LG 14, 16 and 18. Functional candidates in these intervals were inspected. Lastly, taking advantage of available transcriptomic datasets, expression data along berry development were integrated, in order to pinpoint among positional candidates, those differentially expressed across the veraison transition. CONCLUSION:Integration of meta-QTLs analysis on available phenology related QTLs and data from transcriptomic dataset allowed to strongly reduce the number of candidate genes for the genetic control of the veraison transition, prioritizing a list of 272 genes, among which 78 involved in regulation of gene expression, signal transduction or development.
Project description:Grape berry color is an economically important trait that is controlled by two major genes influencing anthocyanin synthesis in the skin. Color is often described qualitatively using six major categories; however, this is a subjective rating that often fails to describe variation within these six classes. To investigate minor genes influencing berry color, image analysis was used to quantify berry color using different color spaces. An image analysis pipeline was developed and utilized to quantify color in a segregating hybrid wine grape population across two years. Images were collected from grape clusters immediately after harvest and segmented by color to determine the red, green, and blue (RGB); hue, saturation, and intensity (HSI); and lightness, red-green, and blue-yellow values (L?a?b?) of berries. QTL analysis identified known major QTL for color on chromosome 2 along with several previously unreported smaller-effect QTL on chromosomes 1, 5, 6, 7, 10, 15, 18, and 19. This study demonstrated the ability of an image analysis phenotyping system to characterize berry color and to more effectively capture variability within a population and identify genetic regions of interest.
Project description:BACKGROUND:Berry color is an important trait in grapes and is mainly determined by the anthocyanin content and composition. To further explore the coloring mechanism of grape berries, the F1 population of Vitis vinifera 'Red Globe'?×?'Muscat Hamburg' was used to map the color locus, and transcriptome analysis was performed to assist in screening candidate genes. RESULTS:A total of 438,407 high-quality single-nucleotide polymorphisms (SNPs) were obtained from whole-genome resequencing (WGS) of the population, and 27,454 SNPs were selected to construct a high-density genetic map. The selected SNPs were clustered into 19 linkage groups (LGs) spanning a genetic distance of 1442.638?cM. Berry color was evaluated by color grade, chromatic aberration, total anthocyanin content and anthocyanin composition. The Pearson correlation coefficients of these phenotypes in 2017 and 2018 were significant at the 0.01 level. The major color locus of MYBA1 and MYBA2 on LG2 was identified, explaining between 26 and 63.6% of all phenotypic variance. Furthermore, 9 additional QTLs with smaller effects were detected on Chr2, Chr4, Chr6, Chr11 and Chr17. Combined with the gene annotation and RNA-seq data, multiple new candidate genes were selected from the above QTLs. CONCLUSION:These results indicated that grape berry color is a quantitative trait controlled by a major color locus and multiple minor loci. Though the major color locus was consistent with previous studies, several minor QTLs and candidate genes associated with grape berry color and anthocyanin accumulation were identified in this study. And the specific regulatory mechanism still needs to be further explored.
Project description:BACKGROUND:Aroma, berry firmness and berry shape are three main quality traits in table grape production, and also the important target traits in grapevine breeding. However, the information about their genetic mechanisms is limited, which results in low accuracy and efficiency of quality breeding in grapevine. Mapping and isolation of quantitative trait locus (QTLs) based on the construction of genetic linkage map is a powerful approach to decipher the genetic determinants of complex quantitative traits. RESULTS:In the present work, a final integrated map consisting of 3411 SLAF markers on 19 linkage groups (LGs) with an average distance of 0.98?cM between adjacent markers was generated using the specific length amplified fragment sequencing (SLAF-seq) technique. A total of 9 significant stable QTLs for Muscat flavor, berry firmness and berry shape were identified on two linkage groups among the hybrids analyzed over three consecutive years from 2016 to 2018. Notably, new stable QTLs for berry firmness and berry shape were found on LG 8 respectively for the first time. Based on biological function and expression profiles of candidate genes in the major QTL regions, 3 genes (VIT_08s0007g00440, VIT_08s0040g02740 and VIT_08s0040g02350) related to berry firmness and 3 genes (VIT_08s0032g01110, VIT_08s0032g01150 and VIT_08s0105g00200) linked to berry shape were highlighted. Overexpression of VIT_08s0032g01110 in transgenic Arabidopsis plants caused the change of pod shape. CONCLUSIONS:A new high-density genetic map with total 3411 markers was constructed with SLAF-seq technique, and thus enabled the detection of narrow interval QTLs for relevant traits in grapevine. VIT_08s0007g00440, VIT_08s0040g02740 and VIT_08s0040g02350 were found to be related to berry firmness, while VIT_08s0032g01110, VIT_08s0032g01150 and VIT_08s0105g00200 were linked to berry shape.
Project description:Deciphering the genetic control of grape berry traits is crucial for optimizing yield, fruit quality, and consumer acceptability. In this study, an association panel of 179 grape genotypes comprising a mixture of ancient cultivars, landraces, and modern varieties collected worldwide were genotyped with genotyping-by-sequencing using a genome-wide association approach based on 32,311 single-nucleotide polymorphism (SNP) markers. Genome-wide efficient mixed-model association was selected as the optimal statistical model based on the results of known control loci of grape berry color traits. Many of the associated SNPs identified in this study were in accordance with the previous QTL analyses using biparental mapping. The grape skin color locus was found to be associated with a mybA transcription factor on chromosome 2. Two strong and distinct association signals associated with berry development periods were found on chromosome 16. Most candidate genes of the interval were highlighted as receptor-like protein kinase. For berry weight, significant association loci were identified on chromosome 18, as previously known, and on chromosome 19 and chromosome 17, as newly mapped. Berry flesh texture was newly located on chromosome 16; candidate genes in the interval were related to calcium. Berry flavor was determined on chromosome 5. Genomic regions were further investigated to reveal candidate genes. In this work, we identified interesting genetic determinants of grape berry-related traits. The identification of the markers closely associated with these berry traits may be useful for grape molecular breeding.
Project description:BACKGROUND: The timing of grape ripening initiation, length of maturation period, berry size and seed content are target traits in viticulture. The availability of early and late ripening varieties is desirable for staggering harvest along growing season, expanding production towards periods when the fruit gets a higher value in the market and ensuring an optimal plant adaptation to climatic and geographic conditions. Berry size determines grape productivity; seedlessness is especially demanded in the table grape market and is negatively correlated to fruit size. These traits result from complex developmental processes modified by genetic, physiological and environmental factors. In order to elucidate their genetic determinism we carried out a quantitative analysis in a 163 individuals-F1 segregating progeny obtained by crossing two table grape cultivars. RESULTS: Molecular linkage maps covering most of the genome (2n = 38 for Vitis vinifera) were generated for each parent. Eighteen pairs of homologous groups were integrated into a consensus map spanning over 1426 cM with 341 markers (mainly microsatellite, AFLP and EST-derived markers) and an average map distance between loci of 4.2 cM. Segregating traits were evaluated in three growing seasons by recording flowering, veraison and ripening dates and by measuring berry size, seed number and weight. QTL (Quantitative Trait Loci) analysis was carried out based on single marker and interval mapping methods. QTLs were identified for all but one of the studied traits, a number of them steadily over more than one year. Clusters of QTLs for different characters were detected, suggesting linkage or pleiotropic effects of loci, as well as regions affecting specific traits. The most interesting QTLs were investigated at the gene level through a bioinformatic analysis of the underlying Pinot noir genomic sequence. CONCLUSION: Our results revealed novel insights into the genetic control of relevant grapevine features. They provide a basis for performing marker-assisted selection and testing the role of specific genes in trait variation.
Project description:Berry firmness is one of the most important quality traits in table grapes. The underlying molecular and genetic mechanisms for berry firmness remain unclear. We constructed a high-density genetic map based on whole-genome resequencing to identify loci associated with berry firmness. The genetic map had 19 linkage groups, including 1662 bin markers (26,039 SNPs), covering 1463.38 cM, and the average inter-marker distance was 0.88 cM. An analysis of berry firmness in the F1 population and both parents for three consecutive years revealed continuous variability in F1, with a distribution close to the normal distribution. Based on the genetic map and phenotypic data, three potentially significant quantitative trait loci (QTLs) related to berry firmness were identified by composite interval mapping. The contribution rate of each QTL ranged from 21.5% to 28.6%. We identified four candidate genes associated with grape firmness, which are related to endoglucanase, abscisic acid (ABA), and transcription factors. A qRT-PCR analysis revealed that the expression of abscisic-aldehyde oxidase-like gene (VIT_18s0041g02410) and endoglucanase 3 gene (VIT_18s0089g00210) in Muscat Hamburg was higher than in Crimson Seedless at the veraison stage, which was consistent with that of parent berry firmness. These results confirmed that VIT_18s0041g02410 and VIT_18s0089g00210 are candidate genes associated with berry firmness.
Project description:BACKGROUND: In grapevine, as in other fruit crops, fruit size and seed content are key components of yield and quality; however, very few Quantitative Trait Loci (QTLs) for berry weight and seed content (number, weight, and dry matter percentage) have been discovered so far. To identify new stable QTLs for marker-assisted selection and candidate gene identification, we performed simultaneous QTL detection in four mapping populations (seeded or seedless) with various genetic backgrounds. RESULTS: For berry weight, we identified five new QTLs, on linkage groups (LGs) 1, 8, 11, 17 and 18, in addition to the known major QTL on LG 18. The QTL with the largest effect explained up to 31% of total variance and was found in two genetically distant populations on LG 17, where it colocalized with a published putative domestication locus. For seed traits, besides the major QTLs on LG 18 previously reported, we found four new QTLs explaining up to 51% of total variance, on LGs 4, 5, 12 and 14. The previously published QTL for seed number on LG 2 was found related in fact to sex. We found colocalizations between seed and berry weight QTLs only for the major QTL on LG 18 in a seedless background, and on LGs 1 and 13 in a seeded background. Candidate genes belonging to the cell number regulator CNR or cytochrome P450 families were found under the berry weight QTLs on LGs 1, 8, and 17. The involvement of these gene families in fruit weight was first described in tomato using a QTL-cloning approach. Several other interesting candidate genes related to cell wall modifications, water import, auxin and ethylene signalling, transcription control, or organ identity were also found under berry weight QTLs. CONCLUSION: We discovered a total of nine new QTLs for berry weight or seed traits in grapevine, thereby increasing more than twofold the number of reliable QTLs for these traits available for marker assisted selection or candidate gene studies. The lack of colocalization between berry and seed QTLs suggests that these traits may be partly dissociated.
Project description:BACKGROUND:Berry size is considered as one of the main selection criteria in table grapes breeding programs, due to the consumer preferences. However, berry size is a complex quantitive trait under polygenic control, and its genetic determination of berry weight is not yet fully understood. The aim of this work was to perform marker discovery using a transcriptomic approach, in order to identify and characterize SNP and InDel markers associated with berry size in table grapes. We used an integrative analysis based on RNA-Seq, SNP/InDel search and validation on table grape segregants and varieties with different genetic backgrounds. RESULTS:Thirty SNPs and eight InDels were identified using a transcriptomic approach (RNA-Seq). These markers were selected from SNP/InDel found among segregants from a Ruby x Sultanina population with contrasting phenotypes for berry size. The set of 38 SNP and InDel markers was distributed in eight chromosomes. Genotype-phenotype association analyses were performed using a set of 13 RxS segregants and 41 table grapes varieties with different genetic backgrounds during three seasons. The results showed several degrees of association of these markers with berry size (10.2 to 30.7%) as other berry-related traits such as length and width. The co-localization of SNP and /or InDel markers and previously reported QTLs and candidate genes associated with berry size were analysed. CONCLUSIONS:We identified a set of informative and transferable SNP and InDel markers associated with berry size. Our results suggest the suitability of SNPs and InDels as candidate markers for berry weight in seedless table grape breeding. The identification of genomic regions associated with berry weight in chromosomes 8, 15 and 17 was achieved with supporting evidence derived from a transcriptome experiment focused on SNP/InDel search, as well as from a QTL-linkage mapping approach. New regions possibly associated with berry weight in chromosomes 3, 6, 9 and 14 were identified.