Genome-wide association study, population structure, and genetic diversity of the tea plant in Guizhou Plateau.

BACKGROUND: Guizhou Plateau, as one of the original centers of tea plant, has a profound multi-ethnic cultural heritage and abundant tea germplasm resources. However, the impact of indigenous community factors on the genetic diversity, population structure and geographical distribution of tea plant is still unclear. RESULTS: Using the genotyping-by-sequencing (GBS) approach, we collected 415 tea plant accessions from the study sites, estimated genetic diversity, developed a core collection, and conducted a genome-wide association study (GWAS) based on 99,363 high-quality single-nucleotide polymorphisms (SNPs). A total of 415 tea accessions were clustered into six populations (GP01, GP02, GP03, GP04, GP05 and GP06), and the results showed that GP04 and GP05 had the highest and lowest genetic diversity (Pi = 0.214 and Pi = 0.145, respectively). Moreover, 136 tea accessions (33%) were selected to construct the core set that can represent the genetic diversity of the whole collection. By analyzing seven significant SNP markers associated with the traits such as the germination period of one bud and two leaves (OTL) and the germination period of one bud and three leaves (OtL), four candidate genes possibly related to OTL and OtL were identified. CONCLUSIONS: This study revealed the impact of indigenous communities on the population structure of 415 tea accessions, indicating the importance of cultural practices for protection and utilization of tea plant genetic resources. Four potential candidate genes associated with the OTL and OtL of tea plant were also identified, which will facilitate genetic research, germplasm conservation, and breeding.

Insights into the genetic architecture of Phytophthora capsici root rot resistance in chile pepper (Capsicum spp.) from multi-locus genome-wide association study.

BACKGROUND: Phytophthora root rot, a major constraint in chile pepper production worldwide, is caused by the soil-borne oomycete, Phytophthora capsici. This study aimed to detect significant regions in the Capsicum genome linked to Phytophthora root rot resistance using a panel consisting of 157 Capsicum spp. genotypes. Multi-locus genome wide association study (GWAS) was conducted using single nucleotide polymorphism (SNP) markers derived from genotyping-by-sequencing (GBS). Individual plants were separately inoculated with P. capsici isolates, 'PWB-185', 'PWB-186', and '6347', at the 4-8 leaf stage and were scored for disease symptoms up to 14-days post-inoculation. Disease scores were used to calculate disease parameters including disease severity index percentage, percent of resistant plants, area under disease progress curve, and estimated marginal means for each genotype. RESULTS: Most of the genotypes displayed root rot symptoms, whereas five accessions were completely resistant to all the isolates and displayed no symptoms of infection. A total of 55,117 SNP markers derived from GBS were used to perform multi-locus GWAS which identified 330 significant SNP markers associated with disease resistance. Of these, 56 SNP markers distributed across all the 12 chromosomes were common across the isolates, indicating association with more durable resistance. Candidate genes including nucleotide-binding site leucine-rich repeat (NBS-LRR), systemic acquired resistance (SAR8.2), and receptor-like kinase (RLKs), were identified within 0.5 Mb of the associated markers. CONCLUSIONS: Results will be used to improve resistance to Phytophthora root rot in chile pepper by the development of Kompetitive allele-specific markers (KASP®) for marker validation, genomewide selection, and marker-assisted breeding.

Consensus genetic linkage map and QTL mapping allow to capture the genomic regions associated with agronomic traits in pearl millet.

MAIN CONCLUSION: A genetic linkage map representing the pearl millet genome was constructed with SNP markers. Major and stable QTL associated with flowering, number of productive tillers, ear head length, and test weight were mapped on chromosomes 1 and 3. Pearl millet (Pennisetum glaucum) is a major cereal and fodder crop in arid and semi-arid regions of Asia and Africa. Agronomic traits are important traits in pearl millet breeding and genetic and environmental factors highly influence them. In the present study, an F9 recombinant inbred line (RIL) population derived from a cross between PT6029 and PT6129 was evaluated for agronomic traits in three environments. Utilizing a genotyping by sequencing approach, a dense genetic map with 993 single nucleotide polymorphism markers covering a total genetic distance of 1035.4 cM was constructed. The average interval between the markers was 1.04 cM, and the seven chromosomes varied from 115.39 to 206.72 cM. Quantitative trait loci (QTL) mapping revealed 35 QTL for seven agronomic traits, and they were distributed on all pearl millet chromosomes. These QTL individually explained 11.35 to 26.71% of the phenotypic variation, with LOD values ranging from 2.74 to 5.80. Notably, four QTL (qDFF1.1, qNPT3.1, qEHL3.1, and qTW1.1) associated with days to fifty percent flowering, the number of productive tillers, ear head length, and test weight were found to be major and stable QTL located on chromosomes 1 and 3. Collectively, our results provide an important base for understanding the genetic architecture of agronomic traits in pearl millet, which is useful for accelerating the genetic gain toward crop improvement.

Uncovering genes involved in pollinator-driven mating system shifts and selfing syndrome evolution in Brassica rapa.

Shifts in pollinator occurrence and their pollen transport effectiveness drive the evolution of mating systems in flowering plants. Understanding the genomic basis of these changes is essential for predicting the persistence of a species under environmental changes. We investigated the genomic changes in Brassica rapa over nine generations of pollination by hoverflies associated with rapid morphological evolution toward the selfing syndrome. We combined a genotyping-by-sequencing (GBS) approach with a genome-wide association study (GWAS) to identify candidate genes, and assessed their functional role in the observed morphological changes by studying mutations of orthologous genes in the model plant Arabidopsis thaliana. We found 31 candidate genes involved in a wide range of functions from DNA/RNA binding to transport. Our functional assessment of orthologous genes in A. thaliana revealed that two of the identified genes in B. rapa are involved in regulating the size of floral organs. We found a protein kinase superfamily protein involved in petal width, an important trait in plant attractiveness to pollinators. Moreover, we found a histone lysine methyltransferase (HKMT) associated with stamen length. Altogether, our study shows that hoverfly pollination leads to rapid evolution toward the selfing syndrome mediated by polygenic changes.

High dispersal ability versus migratory traditions: Fine-scale population structure and post-glacial colonisation in bar-tailed godwits.

In migratory animals, high mobility may reduce population structure through increased dispersal and enable adaptive responses to environmental change, whereas rigid migratory routines predict low dispersal, increased structure, and limited flexibility to respond to change. We explore the global population structure and phylogeographic history of the bar-tailed godwit, Limosa lapponica, a migratory shorebird known for making the longest non-stop flights of any landbird. Using nextRAD sequencing of 14,318 single-nucleotide polymorphisms and scenario-testing in an Approximate Bayesian Computation framework, we infer that bar-tailed godwits existed in two main lineages at the last glacial maximum, when much of their present-day breeding range persisted in a vast, unglaciated Siberian-Beringian refugium, followed by admixture of these lineages in the eastern Palearctic. Subsequently, population structure developed at both longitudinal extremes: in the east, a genetic cline exists across latitude in the Alaska breeding range of subspecies L. l. baueri; in the west, one lineage diversified into three extant subspecies L. l. lapponica, taymyrensis, and yamalensis, the former two of which migrate through previously glaciated western Europe. In the global range of this long-distance migrant, we found evidence of both (1) fidelity to rigid behavioural routines promoting fine-scale geographic population structure (in the east) and (2) flexibility to colonise recently available migratory flyways and non-breeding areas (in the west). Our results suggest that cultural traditions in highly mobile vertebrates can override the expected effects of high dispersal ability on population structure, and provide insights for the evolution and flexibility of some of the world's longest migrations.

The impact of geographic isolation and host shifts on population divergence of the rare cicada Subpsaltria yangi.

The contributions of divergent selection and spatial isolation to population divergence are among the main focuses of evolutionary biology. Here we employed integrated methods to explore genomic divergence, demographic history and calling-song differentiation in the cicada Subpsaltria yangi, and compared the genotype and calling-song phenotype of different populations occurring in distinct habitats. Our results indicate that this species comprises four main lineages with unique sets of haplotypes and calling-song structure, which are distinctly associated with geographic isolation and habitats. The populations occurring on the Loess Plateau underwent substantial expansion at âˆ¼0.130-0.115 Ma during the Last Interglacial. Geographic distance and host shift between pairs of populations predict genomic divergence, with geographic distance and acoustical signal together explaining > 60% of the divergence among populations. Differences in calling songs could reflect adaptation of populations to novel environments with different host plants, habitats and predators, which may have resulted from neutral divergence at the molecular level followed by natural selection. Geomorphic barriers and climate oscillations associated with Pleistocene glaciation may have been primary factors in shaping the population genetic structure of this species. Ultimately this may couple with a host shift in leading toward allopatric speciation in S. yangi, i.e., isolation by distance. Our findings improve understanding of divergence in allopatry of herbivorous insects, and may inform future studies on the molecular mechanisms underlying the association between genetic/phenotypic changes and adaptation of insects to novel niches and host plants.

Comparison of the genetic basis of salt tolerance at germination, seedling, and reproductive stages in an introgression line population of rice.

BACKGROUND: Salinity is a major limitation for rice farming due to climate change. Since salt stress adversely impact rice plants at germination, seedling, and reproductive stages resulting in poor crop establishment and reduced grain yield, enhancing salt tolerance at these vulnerable growth stages will enhance rice productivity in salinity prone areas. METHODS AND RESULTS: An introgression line (ILs) population from a cross between a high yielding cultivar 'Cheniere' and a salt tolerant donor 'TCCP' was evaluated to map quantitative trait loci (QTLs) for traits associated with salt tolerance at germination, seedling, and reproductive stages. Using a genotyping-by-sequencing based high density SNP linkage map, a total of 7, 16, and 30 QTLs were identified for five germination traits, seven seedling traits, and ten reproductive traits, respectively. There was overlapping of QTLs for some traits at different stages indicating the pleiotropic effects of these QTLs or clustering of linked genes. Candidate genes identified for salt tolerance were OsSDIR1 and SERF for the seedling stage, WRKY55 and OsUBC for the reproductive stage, and MYB family transcription factors for all three stages. Gene ontology analysis revealed significant GO terms related to nucleotide binding, protein binding, protein kinase activity, antiporter activity, active transmembrane transporter activity, calcium-binding protein, and F- box protein interaction domain containing protein. CONCLUSIONS: The colocalized QTLs for traits at different growth stages would be helpful to improve multiple traits simultaneously using marker-assisted selection. The salt tolerant ILs have the potential to be released as varieties or as pre-breeding lines for developing salt tolerant rice varieties.

Unveiling population structure and selection signatures of riverine and genetically improved rohu, Labeo rohita using genome wide SNPs.

BACKGROUND: Captive breeding, along with artificial selection can significantly impact population structure by influencing allele frequencies and driving populations towards specific adaptation. Selective sweeps are powerful forces in shaping genetic variation within populations and can drive rapid spread of beneficial alleles while simultaneously reducing genetic diversity in localized regions of the genome. The present work was undertaken to assess the genetic structure and consequences of artificial selection in 10th generation of genetically improved rohu by comparing with wild populations. METHODS AND RESULTS: The present study used 11,022 high-quality genome wide SNPs to compare the population genetic structure and signatures of selection between Jayanti rohu population and its wild counterpart. Outlier analysis revealed presence of 14 adaptive SNPs, out of which 5 were classified to be under decisive selection pressure. Notably, Jayanti rohu (JR) displayed 297 private alleles exclusive to its population. Chromosomes 7 and 16 emerged as potential hotspots containing a majority of the identified SNPs. Structure and principal component analysis revealed two distinct clusters, effectively distinguishing the JR and wild rohu populations. Phylogenetic analysis indicated a separate cluster of JR population distant from wild groups. CONCLUSION: The results of present study shall help in elucidating patterns of genetic variation and characterizing selection signatures associated with captive bred and natural populations of rohu. The genomic resources generated through this work shall be helpful in improving the traceability of selectively bred germplasm for developing future strategies of genetic management.

Characterization of Quantitative Trait Loci for Leaf Rust Resistance from CI 13227 in Three Winter Wheat Populations.

Leaf rust is a widespread foliar wheat disease causing substantial yield losses worldwide. Slow rusting is "adult plant" resistance that significantly slows epidemic development and thereby reduces yield loss. Wheat accession CI 13227 was previously characterized as having slow-rusting resistance. To validate the quantitative trait loci (QTLs) and develop diagnostic markers for slow rusting resistance in CI 13227, a new population of recombinant inbred lines of CI 13227 × Everest was evaluated for latent period, final severity, area under the disease progress curve, and infection type in greenhouses and genotyped using genotyping-by-sequencing. Four QTLs were identified on chromosome arms 2BL, 2DS, 3BS, and 7BL, explaining 6.82 to 28.45% of the phenotypic variance for these traits. Seven kompetitive allele-specific polymorphism markers previously reported to be linked to the QTLs in two other CI 13227 populations were validated. In addition, the previously reported QLr.hwwg-7AL was remapped to 2BL (renamed QLr.hwwg-2BL) after adding new markers in this study. Phenotypic data showed that the recombinant inbred lines harboring two or three of the QTLs had a significantly longer latent period. QLr.hwwg-2DS on 2DS showed a major effect on all rust resistance traits and was finely mapped to a 2.7-Mb interval by two newly developed flanking markers from exome capture. Three disease-resistance genes and two transporter genes were identified as the putative candidates for QLr.hwwg-2DS. The validated QTLs can be used as slow-rusting resistance resources, and the markers developed in this study will be useful for marker-assisted selection.

Contrasting genetic signal of recolonization after rainforest fragmentation in African trees with different dispersal abilities.

Although today the forest cover is continuous in Central Africa, this may have not always been the case, as the scarce fossil record in this region suggests that arid conditions might have significantly reduced tree density during the ice ages. Our aim was to investigate whether the dry ice age periods left a genetic signature on tree species that can be used to infer the date of the past fragmentation of the rainforest. We sequenced reduced representation libraries of 182 samples representing five widespread legume trees and seven outgroups. Phylogenetic analyses identified an early divergent lineage for all species in West Africa (Upper Guinea) and two clades in Central Africa: Lower Guinea-North and Lower Guinea-South. As the structure separating the Northern and Southern clades-congruent across species-cannot be explained by geographic barriers, we tested other hypotheses with demographic model testing using δαδι. The best estimates indicate that the two clades split between the Upper Pliocene and the Pleistocene, a date compatible with forest fragmentation driven by ice age climatic oscillations. Furthermore, we found remarkably older split dates for the shade-tolerant tree species with nonassisted seed dispersal than for light-demanding species with long-distance wind-dispersed seeds. Different recolonization abilities after recurrent cycles of forest fragmentation seem to explain why species with long-distance dispersal show more recent genetic admixture between the two clades than species with limited seed dispersal. Despite their old history, our results depict the African rainforests as a dynamic biome where tree species have expanded relatively recently after the last glaciation.

Imputation strategies for genomic prediction using nanopore sequencing.

BACKGROUND: Genomic prediction describes the use of SNP genotypes to predict complex traits and has been widely applied in humans and agricultural species. Genotyping-by-sequencing, a method which uses low-coverage sequence data paired with genotype imputation, is becoming an increasingly popular SNP genotyping method for genomic prediction. The development of Oxford Nanopore Technologies' (ONT) MinION sequencer has now made genotyping-by-sequencing portable and rapid. Here we evaluate the speed and accuracy of genomic predictions using low-coverage ONT sequence data in a population of cattle using four imputation approaches. We also investigate the effect of SNP reference panel size on imputation performance. RESULTS: SNP array genotypes and ONT sequence data for 62 beef heifers were used to calculate genomic estimated breeding values (GEBVs) from 641 k SNP for four traits. GEBV accuracy was much higher when genome-wide flanking SNP from sequence data were used to help impute the 641 k panel used for genomic predictions. Using the imputation package QUILT, correlations between ONT and low-density SNP array genomic breeding values were greater than 0.91 and up to 0.97 for sequencing coverages as low as 0.1 × using a reference panel of 48 million SNP. Imputation time was significantly reduced by decreasing the number of flanking sequence SNP used in imputation for all methods. When compared to high-density SNP arrays, genotyping accuracy and genomic breeding value correlations at 0.5 × coverage were also found to be higher than those imputed from low-density arrays. CONCLUSIONS: Here we demonstrated accurate genomic prediction is possible with ONT sequence data from sequencing coverages as low as 0.1 × , and imputation time can be as short as 10 min per sample. We also demonstrate that in this population, genotyping-by-sequencing at 0.1 × coverage can be more accurate than imputation from low-density SNP arrays.

Genome wide association study of genes controlling resistance to Didymella rabiei Pathotype IV through genotyping by sequencing in chickpeas (Cicer arietinum).

Ascochyta blight (AB) is a major disease in chickpeas (Cicer arietinum L.) that can cause a yield loss of up to 100%. Chickpea germplasm collections at the center of origin offer great potential to discover novel sources of resistance to pests and diseases. Herein, 189 Cicer arietinum samples were genotyped via genotyping by sequencing. This chickpea collection was phenotyped for resistance to an aggressive Turkish Didymella rabiei Pathotype IV isolate. Genome-wide association studies based on different models revealed 19 single nucleotide polymorphism (SNP) associations on chromosomes 1, 2, 3, 4, 7, and 8. Although eight of these SNPs have been previously reported, to the best of our knowledge, the remaining ten were associated with AB resistance for the first time. The regions identified in this study can be addressed in future studies to reveal the genetic mechanism underlying AB resistance and can also be utilized in chickpea breeding programs to improve AB resistance in new chickpea varieties.

Patterns of the Predicted Mutation Burden in 19,778 Domesticated Barley Accessions Conserved Ex Situ.

Long-term conservation of more than 7 million plant germplasm accessions in 1750 genebanks worldwide is a challenging mission. The extent of deleterious mutations present in conserved germplasm and the genetic risk associated with accumulative mutations are largely unknown. This study took advantage of published barley genomic data to predict sample-wise mutation burdens for 19,778 domesticated barley (Hordeum vulgare L.) accessions conserved ex situ. It was found that the conserved germplasm harbored 407 deleterious mutations and 337 (or 82%) identified deleterious alleles were present in 20 (or 0.1%) or fewer barley accessions. Analysis of the predicted mutation burdens revealed significant differences in mutation burden for several groups of barley germplasm (landrace > cultivar (or higher burden estimate in landrace than in cultivar); winter barley > spring barley; six-rowed barley > two-rowed barley; and 1000-accession core collection > non-core germplasm). Significant differences in burden estimate were also found among seven major geographical regions. The sample-wise predicted mutation burdens were positively correlated with the estimates of sample average pairwise genetic difference. These findings are significant for barley germplasm management and utilization and for a better understanding of the genetic risk in conserved plant germplasm.

Genetic Diversity and Genome-Wide Association Analysis of the Hulled/Naked Trait in a Barley Collection from Shanghai Agricultural Gene Bank.

Barley is one of the most important cereal crops in the world, and its value as a food is constantly being revealed, so the research into and the use of barley germplasm are very important for global food security. Although a large number of barley germplasm samples have been collected globally, their specific genetic compositions are not well understood, and in many cases their origins are even disputed. In this study, 183 barley germplasm samples from the Shanghai Agricultural Gene Bank were genotyped using genotyping-by-sequencing (GBS) technology, SNPs were identified and their genetic parameters were estimated, principal component analysis (PCA) was preformed, and the phylogenetic tree and population structure of the samples were also analyzed. In addition, a genome-wide association study (GWAS) was carried out for the hulled/naked grain trait, and a KASP marker was developed using an associated SNP. The results showed that a total of 181,906 SNPs were identified, and these barley germplasm samples could be roughly divided into three categories according to the phylogenetic analysis, which was generally consistent with the classification of the traits of row type and hulled/naked grain. Population structure analysis showed that the whole barley population could be divided into four sub-populations (SPs), the main difference from previous classifications being that the two-rowed and the hulled genotypes were sub-divided into two SPs. The GWAS analysis of the hulled/naked trait showed that many associated loci were unrelated to the Nud/nud locus, indicating that there might be new loci controlling the trait. A KASP marker was developed for one exon-type SNP on chromosome 7. Genotyping based on the KASP assay was consistent with that based on SNPs, indicating that the gene of this locus might be associated with the hulled/naked trait. The above work not only lays a good foundation for the future utilization of this barley germplasm population but it provides new loci and candidate genes for the hulled/naked trait.

Genetic analysis of yield components in buckwheat using high-throughput sequencing analysis and wild resource populations.

Fagopyrum tataricum, an important medicinal and edible crop, possesses significant agricultural and economic value. However, the development of buckwheat varieties and yields has been hindered by the delayed breeding progress despite the abundant material resources in China. Current research indicates that quantitative trait loci (QTLs) play a crucial role in controlling plant seed type and yield. To address these limitations, this study constructed recombinant inbred lines (RILs) utilizing both cultivated species and wild buckwheat as raw materials. In total, 84,521 Single Nucleotide Polymorphism (SNP) markers were identified through Genotyping-by-Sequencing (GBS) technology, and high-resolution and high-density SNP genetic maps were developed, which had significant value for QTL mapping, gene cloning and comparative mapping of buckwheat. In this study, we successfully identified 5 QTLs related to thousand grain weight (TGW), 9 for grain length (GL), and 1 for grain width (GW) by combining seed type and TGW data from 202 RIL populations in four different environments, within which one co-located QTL for TGW were discovered on the first chromosome. Transcriptome analysis during different grain development stages revealed 59 significant expression differences between the two materials, which can serve as candidate genes for further investigation into the regulation of grain weight and yield enhancement. The mapped major loci controlling TGW, GL and GW will be valuable for gene cloning and reveal the mechanism underlying grain development and marker-assisted selection in Tartary buckwheat.

Insights into breeding history, hotspot regions of selection, and untapped allelic diversity for bread wheat breeding.

Breeding has increasingly altered the genetics of crop plants since the domestication of their wild progenitors. It is postulated that the genetic diversity of elite wheat breeding pools is too narrow to cope with future challenges. In contrast, plant genetic resources (PGRs) of wheat stored in genebanks are valuable sources of unexploited genetic diversity. Therefore, to ensure breeding progress in the future, it is of prime importance to identify the useful allelic diversity available in PGRs and to transfer it into elite breeding pools. Here, a diverse collection consisting of modern winter wheat cultivars and genebank accessions was investigated based on reduced-representation genomic sequencing and an iSelect single nucleotide polymorphism (SNP) chip array. Analyses of these datasets provided detailed insights into population structure, levels of genetic diversity, sources of new allelic diversity, and genomic regions affected by breeding activities. We identified 57 regions representing genomic signatures of selection and 827 regions representing private alleles associated exclusively with genebank accessions. The presence of known functional wheat genes, quantitative trait loci, and large chromosomal modifications, i.e., introgressions from wheat wild relatives, provided initial evidence for putative traits associated within these identified regions. These findings were supported by the results of ontology enrichment analyses. The results reported here will stimulate further research and promote breeding in the future by allowing for the targeted introduction of novel allelic diversity into elite wheat breeding pools.

Sorghum Association Panel whole-genome sequencing establishes cornerstone resource for dissecting genomic diversity.

Association mapping panels represent foundational resources for understanding the genetic basis of phenotypic diversity and serve to advance plant breeding by exploring genetic variation across diverse accessions. We report the whole-genome sequencing (WGS) of 400 sorghum (Sorghum bicolor (L.) Moench) accessions from the Sorghum Association Panel (SAP) at an average coverage of 38× (25-72×), enabling the development of a high-density genomic marker set of 43 983 694 variants including single-nucleotide polymorphisms (approximately 38 million), insertions/deletions (indels) (approximately 5 million), and copy number variants (CNVs) (approximately 170 000). We observe slightly more deletions among indels and a much higher prevalence of deletions among CNVs compared to insertions. This new marker set enabled the identification of several novel putative genomic associations for plant height and tannin content, which were not identified when using previous lower-density marker sets. WGS identified and scored variants in 5-kb bins where available genotyping-by-sequencing (GBS) data captured no variants, with half of all bins in the genome falling into this category. The predictive ability of genomic best unbiased linear predictor (GBLUP) models was increased by an average of 30% by using WGS markers rather than GBS markers. We identified 18 selection peaks across subpopulations that formed due to evolutionary divergence during domestication, and we found six Fst peaks resulting from comparisons between converted lines and breeding lines within the SAP that were distinct from the peaks associated with historic selection. This population has served and continues to serve as a significant public resource for sorghum research and demonstrates the value of improving upon existing genomic resources.

Large-scale analysis of sheep rumen metagenome profiles captured by reduced representation sequencing reveals individual profiles are influenced by the environment and genetics of the host.

BACKGROUND: Producing animal protein while reducing the animal's impact on the environment, e.g., through improved feed efficiency and lowered methane emissions, has gained interest in recent years. Genetic selection is one possible path to reduce the environmental impact of livestock production, but these traits are difficult and expensive to measure on many animals. The rumen microbiome may serve as a proxy for these traits due to its role in feed digestion. Restriction enzyme-reduced representation sequencing (RE-RRS) is a high-throughput and cost-effective approach to rumen metagenome profiling, but the systematic (e.g., sequencing) and biological factors influencing the resulting reference based (RB) and reference free (RF) profiles need to be explored before widespread industry adoption is possible. RESULTS: Metagenome profiles were generated by RE-RRS of 4,479 rumen samples collected from 1,708 sheep, and assigned to eight groups based on diet, age, time off feed, and country (New Zealand or Australia) at the time of sample collection. Systematic effects were found to have minimal influence on metagenome profiles. Diet was a major driver of differences between samples, followed by time off feed, then age of the sheep. The RF approach resulted in more reads being assigned per sample and afforded greater resolution when distinguishing between groups than the RB approach. Normalizing relative abundances within the sampling Cohort abolished structures related to age, diet, and time off feed, allowing a clear signal based on methane emissions to be elucidated. Genus-level abundances of rumen microbes showed low-to-moderate heritability and repeatability and were consistent between diets. CONCLUSIONS: Variation in rumen metagenomic profiles was influenced by diet, age, time off feed and genetics. Not accounting for environmental factors may limit the ability to associate the profile with traits of interest. However, these differences can be accounted for by adjusting for Cohort effects, revealing robust biological signals. The abundances of some genera were consistently heritable and repeatable across different environments, suggesting that metagenomic profiles could be used to predict an individual's future performance, or performance of its offspring, in a range of environments. These results highlight the potential of using rumen metagenomic profiles for selection purposes in a practical, agricultural setting.

Restriction site-associated DNA sequencing technologies as an alternative to low-density SNP chips for genomic selection: a simulation study in layer chickens.

BACKGROUND: To reduce the cost of genomic selection, a low-density (LD) single nucleotide polymorphism (SNP) chip can be used in combination with imputation for genotyping selection candidates instead of using a high-density (HD) SNP chip. Next-generation sequencing (NGS) techniques have been increasingly used in livestock species but remain expensive for routine use for genomic selection. An alternative and cost-efficient solution is to use restriction site-associated DNA sequencing (RADseq) techniques to sequence only a fraction of the genome using restriction enzymes. From this perspective, use of RADseq techniques followed by an imputation step on HD chip as alternatives to LD chips for genomic selection was studied in a pure layer line. RESULTS: Genome reduction and sequencing fragments were identified on reference genome using four restriction enzymes (EcoRI, TaqI, AvaII and PstI) and a double-digest RADseq (ddRADseq) method (TaqI-PstI). The SNPs contained in these fragments were detected from the 20X sequence data of the individuals in our population. Imputation accuracy on HD chip with these genotypes was assessed as the mean correlation between true and imputed genotypes. Several production traits were evaluated using single-step GBLUP methodology. The impact of imputation errors on the ranking of the selection candidates was assessed by comparing a genomic evaluation based on ancestry using true HD or imputed HD genotyping. The relative accuracy of genomic estimated breeding values (GEBVs) was investigated by considering the GEBVs estimated on offspring as a reference. With AvaII or PstI and ddRADseq with TaqI and PstI, more than 10 K SNPs were detected in common with the HD SNP chip, resulting in an imputation accuracy greater than 0.97. The impact of imputation errors on genomic evaluation of the breeders was reduced, with a Spearman correlation greater than 0.99. Finally, the relative accuracy of GEBVs was equivalent. CONCLUSIONS: RADseq approaches can be interesting alternatives to low-density SNP chips for genomic selection. With more than 10 K SNPs in common with the SNPs of the HD SNP chip, good imputation and genomic evaluation results can be obtained. However, with real data, heterogeneity between individuals with missing data must be considered.

Genetic diversity and genome-wide association study of 13 agronomic traits in 977 Beta vulgaris L. germplasms.

BACKGROUND: Sugar beet (Beta vulgaris L.) is an economically essential sugar crop worldwide. Its agronomic traits are highly diverse and phenotypically plastic, influencing taproot yield and quality. The National Beet Medium-term Gene Bank in China maintains more than 1700 beet germplasms with diverse countries of origin. However, it lacks detailed genetic background associated with morphological variability and diversity. RESULTS: Here, a comprehensive genome-wide association study (GWAS) of 13 agronomic traits was conducted in a panel of 977 sugar beet accessions. Almost all phenotypic traits exhibited wide genetic diversity and high coefficient of variation (CV). A total of 170,750 high-quality single-nucleotide polymorphisms (SNPs) were obtained using the genotyping-by-sequencing (GBS). Neighbour-joining phylogenetic analysis, principal component analysis, population structure and kinship showed no obvious relationships among these genotypes based on subgroups or regional sources. GWAS was carried out using a mixed linear model, and 159 significant associations were detected for these traits. Within the 25 kb linkage disequilibrium decay of the associated markers, NRT1/PTR FAMILY 6.3 (BVRB_5g097760); nudix hydrolase 15 (BVRB_8g182070) and TRANSPORT INHIBITOR RESPONSE 1 (BVRB_8g181550); transcription factor MYB77 (BVRB_2g023500); and ethylene-responsive transcription factor ERF014 (BVRB_1g000090) were predicted to be strongly associated with the taproot traits of root groove depth (RGD); root shape (RS); crown size (CS); and flesh colour (FC), respectively. For the aboveground traits, UDP-glycosyltransferase 79B6 (BVRB_9g223780) and NAC domain-containing protein 7 (BVRB_5g097990); F-box protein At1g10780 (BVRB_6g140760); phosphate transporter PHO1 (BVRB_3g048660); F-box protein CPR1 (BVRB_8g181140); and transcription factor MYB77 (BVRB_2g023500) and alcohol acyltransferase 9 (BVRB_2g023460) might be associated with the hypocotyl colour (HC); plant type (PT); petiole length (PL); cotyledon size (C); and fascicled leaf type (FLT) of sugar beet, respectively. AP-2 complex subunit mu (BVRB_5g106130), trihelix transcription factor ASIL2 (BVRB_2g041790) and late embryogenesis abundant protein 18 (BVRB_5g106150) might be involved in pollen quantity (PQ) variation. The candidate genes extensively participated in hormone response, nitrogen and phosphorus transportation, secondary metabolism, fertilization and embryo maturation. CONCLUSIONS: The genetic basis of agronomical traits is complicated in heterozygous diploid sugar beet. The putative valuable genes found in this study will help further elucidate the molecular mechanism of each phenotypic trait for beet breeding.