Dissecting genetic architecture of grape proanthocyanidin composition through quantitative trait locus mapping

<p>Abstract</p> <p>Background</p> <p>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.</p> <p>Results</p> <p>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 (<it>VvLAR1</it>, <it>VvMYBPA2</it>, <it>VvCHI1</it>, <it>VvMYBPA1</it>) showed at least one significant association with PA variables, especially <it>VvLAR1 </it>revealed as of great interest for further functional investigation. Some SNP-phenotype associations were observed only in the diversity panel.</p> <p>Conclusions</p> <p>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.</p

Patterns of sequence polymorphism in the fleshless berry locus in cultivated and wild Vitis vinifera accessions

<p>Abstract</p> <p>Background</p> <p>Unlike in tomato, little is known about the genetic and molecular control of fleshy fruit development of perennial fruit trees like grapevine (<it>Vitis vinifera </it>L.). Here we present the study of the sequence polymorphism in a 1 Mb grapevine genome region at the top of chromosome 18 carrying the <it>fleshless berry </it>mutation (<it>flb</it>) in order, first to identify SNP markers closely linked to the gene and second to search for possible signatures of domestication.</p> <p>Results</p> <p>In total, 62 regions (17 SSR, 3 SNP, 1 CAPS and 41 re-sequenced gene fragments) were scanned for polymorphism along a 3.4 Mb interval (85,127-3,506,060 bp) at the top of the chromosome 18, in both <it>V. vinifera cv</it>. Chardonnay and a genotype carrying the <it>flb </it>mutation, <it>V. vinifera cv</it>. Ugni Blanc mutant. A nearly complete homozygosity in Ugni Blanc (wild and mutant forms) and an expected high level of heterozygosity in Chardonnay were revealed. Experiments using qPCR and BAC FISH confirmed the observed homozygosity. Under the assumption that <it>flb </it>could be one of the genes involved into the domestication syndrome of grapevine, we sequenced 69 gene fragments, spread over the <it>flb </it>region, representing 48,874 bp in a highly diverse set of cultivated and wild <it>V. vinifera </it>genotypes, to identify possible signatures of domestication in the cultivated <it>V. vinifera </it>compartment. We identified eight gene fragments presenting a significant deviation from neutrality of the Tajima's D parameter in the cultivated pool. One of these also showed higher nucleotide diversity in the wild compartments than in the cultivated compartments. In addition, SNPs significantly associated to berry weight variation were identified in the <it>flb </it>region.</p> <p>Conclusions</p> <p>We observed the occurrence of a large homozygous region in a non-repetitive region of the grapevine otherwise highly-heterozygous genome and propose a hypothesis for its formation. We demonstrated the feasibility to apply BAC FISH on the very small grapevine chromosomes and provided a specific probe for the identification of chromosome 18 on a cytogenetic map. We evidenced genes showing putative signatures of selection and SNPs significantly associated with berry weight variation in the <it>flb </it>region. In addition, we provided to the community 554 SNPs at the top of chromosome 18 for the development of a genotyping chip for future fine mapping of the <it>flb </it>gene in a F2 population when available.</p

Analyse et exploitation de séquences Sanger

École thématiqu

Variation in crossing-over rates across chromosome 4 of Arabidopsis thaliana reveals the presence of meiotic recombination "hot spots"

Crossover (CO) is a key process for the accurate segregation of homologous chromosomes during the first meiotic division. In most eukaryotes, meiotic recombination is not homogeneous along the chromosomes, suggesting a tight control of the location of recombination events. We genotyped 71 single nucleotide polymorphisms (SNPs) covering the entire chromosome 4 of Arabidopsis thaliana on 702 F2 plants, representing 1404 meioses and allowing the detection of 1171 COs, to study CO localization in a higher plant. The genetic recombination rates varied along the chromosome from 0 cM/Mb near the centromere to 20 cM/Mb on the short arm next to the NOR region, with a chromosome average of 4.6 cM/Mb. Principal component analysis showed that CO rates negatively correlate with the G+C content (P =3x10(-4)), in contrast to that reported in other eukaryotes. COs also significantly correlate with the density of single repeats and the CpG ratio, but not with genes, pseudogenes, transposable elements, or dispersed repeats. Chromosome 4 has, on average, 1.6 COs per meiosis, and these COs are subjected to interference. A detailed analysis of several regions having high CO rates revealed "hot spots" of meiotic recombination contained in small fragments of a few kilobases. Both the intensity and the density of these hot spots explain the variation of CO rates along the chromosome

Long Reads for Long-Term Goals : A Genome Resource for Lavander Genetic Improvement

International audienceLavender (Lavandula angustifolia L.) is both a patrimonial and economically important species, used for its essential oils in cosmetics and perfumery products. Naturally adapted to poor soil and dry environment, lavender represents a promising culture in the context of climate change. However, increasing pathogenic pressure related to climate change challenges present varieties yields and quality of essential oils. The cultivar 'Maillette', that is most-valued for the quality of its essential oil, is widely used in genetic improvement programs in France. Useful genomic resources available for these programs remain nevertheless scarce, despite a short-reads assembled genome available. To support the emerging programs for genetic improvement in lavender, we generated genomic resources based on long-reads sequences, in complement with Illumina short-reads sequences

BSA-Seq : un outil efficace pour caractériser des loci impliqués dans la résistance à la rouille foliaire du peuplier.

International audienceThe efficiency of the Bulk Segregant Analysis (BSA) had clearly been demonstrated to detect genomic regions and genes involved in diverse traits. It allows large experiments reducing the cost and time and preserving the power of full individual's population analysis. These past few years the combination of BSA and New Generation Sequence (NGS) data (BSA-Seq) gave a new accuracy and depth to the discovery on many traits of interest, mainly on crop and model species. In our study, we applied the BSA-Seq to narrow down Populus genomic regions involved in the resistance to Melampsora larici populina (Mlp) leaf rust. We worked on the 1417 progenies derived from an interspecific cross Populus deltoides clone 73028-62 (Pd) x Populus trichocarpa clone 101-74 (Pt) in which segregates qualitative and quantitative Mlp resistances. Four bulks were constituted based on (1) the phenotypes for uredinias size (bulk1: large, bulk2 and bulk3: intermediate, bulk4: small) and (2) the Pt genotypes at the RUS locus governing the uredinia size (bulk1 and bulk3 : [RUS], bulk2 and bulk4 : [rUS]). We used independently the soft masked genomes of Populus trichocarpa Nisqually v3.0 (Ptv3) and Populus deltoides v2.0 (Pdv2) as references to map parents and bulks Illumina reads with the BWAmem/0.7.15 suite and to detect DNA variations with Freebayes/0.9.21. For each common variant position between parents and bulks, we tracked the specific alleles of Pt and/or Pd in the bulks. Respectively from Ptv3 and Pdv2, we identified 13 and 11 regions. We first evaluated the strategy retrieving the previously finally mapped RUS gene, designed new genomic markers from Ptv3 to better characterize this locus and performed the in silico validation. Then we proceed with it to fine map the other regions. Most of them co-locate with QTL and could explain the resistance to Mlp. So we demonstrated that, in our context, even if the reference genome is different to the studied genomes, BSA-Seq allowed us to detect quickly and costly, regions which may be involved in a complex trait and improve the fine mapping of a major gene. We think it can be a promising method on a high heterozygous diploid genome as Poplar, to decipher complex trait. Next step is to identify candidate genes within the regions and better describe the mechanisms of resistance

Oxford Nanopore and Bionano Genomics technologies evaluation for plant structural variation detection

International audienceAbstract Background Structural Variations (SVs) are genomic rearrangements derived from duplication, deletion, insertion, inversion, and translocation events. In the past, SVs detection was limited to cytological approaches, then to Next-Generation Sequencing (NGS) short reads and partitioned assemblies. Nowadays, technologies such as DNA long read sequencing and optical mapping have revolutionized the understanding of SVs in genomes, due to the enhancement of the power of SVs detection. This study aims to investigate performance of two techniques, 1) long-read sequencing obtained with the MinION device (Oxford Nanopore Technologies) and 2) optical mapping obtained with Saphyr device (Bionano Genomics) to detect and characterize SVs in the genomes of the two ecotypes of Arabidopsis thaliana, Columbia-0 (Col-0) and Landsberg erecta 1 (L er -1). Results We described the SVs detected from the alignment of the best ONT assembly and DLE-1 optical maps of A. thaliana L er -1 against the public reference genome Col-0 TAIR10.1. After filtering (SV > 1 kb), 1184 and 591 L er -1 SVs were retained from ONT and Bionano technologies respectively. A total of 948 L er -1 ONT SVs (80.1%) corresponded to 563 Bionano SVs (95.3%) leading to 563 common locations. The specific locations were scrutinized to assess improvement in SV detection by either technology. The ONT SVs were mostly detected near TE and gene features, and resistance genes seemed particularly impacted. Conclusions Structural variations linked to ONT sequencing error were removed and false positives limited, with high quality Bionano SVs being conserved. When compared with the Col-0 TAIR10.1 reference genome, most of the detected SVs discovered by both technologies were found in the same locations. ONT assembly sequence leads to more specific SVs than Bionano one, the latter being more efficient to characterize large SVs. Even if both technologies are complementary approaches, ONT data appears to be more adapted to large scale populations studies, while Bionano performs better in improving assembly and describing specificity of a genome compared to a reference

BSA-Seq : an efficient method to decipher a complex trait on Poplar, a highly heterozygous diploid genome

The efficiency of the Bulk Segregant Analysis (BSA) had clearly been demonstrated to detect genomic regions and genes involved in diverse traits. It allows large experiments reducing the cost and time and preserving the power of full individual's population analysis. These past few years the combination of BSA and New Generation Sequence (NGS) data (BSA-Seq) gave a new accuracy and depth to the discovery on many traits of interest, mainly on crop and model species. In our study, we applied the BSA-Seq in a heterozygous and diploid genome context. We worked on the progenies derived from an interspecific cross Populus deltoides x Populus trichocarpa in which segregates the resistance to Melampsora larici populina (Mlp) leaf rust. We detected DNA variations with freebayes/0.9.21 and the soft masked genome of Populus trichocarpa Nisqually v3.0 as reference. Comparing DNA variations in between parents and bulks we obtained 27 regions or Quantitative Trait Loci based on NGS analysis (QTL-Seq) which could explain the resistance to Mlp. We first evaluated the strategy retrieving a previously cloned Mlp resistance gene governing the uredinia size in Populus trichocarpa clone 101-74 (RUS). Then we identified genomic markers which should better characterize this locus. So we demonstrated that, in our context, BSA-Seq allows us to improve the fine mapping of a major gene. We think it can be a promising method on a high heterozygous diploid genome as Poplar, to decipher complex trait. Next step is to proceed with it to fine map the other QTL-Seq