A large nested association mapping population for breeding and quantitative trait locus mapping in Ethiopian durum wheat

The Ethiopian plateau hosts thousands of durum wheat (Triticum turgidum subsp. durum) farmer varieties (FV) with high adaptability and breeding potential. To harness their unique allelic diversity, we produced a large nested association mapping (NAM) population intercrossing fifty Ethiopian FVs with an international elite durum wheat variety (Asassa). The Ethiopian NAM population (EtNAM) is composed of fifty interconnected bi‐parental families, totalling 6280 recombinant inbred lines (RILs) that represent both a powerful quantitative trait loci (QTL) mapping tool, and a large pre‐breeding panel. Here, we discuss the molecular and phenotypic diversity of the EtNAM founder lines, then we use an array featuring 13 000 single nucleotide polymorphisms (SNPs) to characterize a subset of 1200 EtNAM RILs from 12 families. Finally, we test the usefulness of the population by mapping phenology traits and plant height using a genome wide association (GWA) approach. EtNAM RILs showed high allelic variation and a genetic makeup combining genetic diversity from Ethiopian FVs with the international durum wheat allele pool. EtNAM SNP data were projected on the fully sequenced AB genome of wild emmer wheat, and were used to estimate pairwise linkage disequilibrium (LD) measures that reported an LD decay distance of 7.4 Mb on average, and balanced founder contributions across EtNAM families. GWA analyses identified 11 genomic loci individually affecting up to 3 days in flowering time and more than 1.6 cm in height. We argue that the EtNAM is a powerful tool to support the production of new durum wheat varieties targeting local and global agriculture.     

High‐density genotyping of the A.E. Watkins Collection of hexaploid landraces identifies a large molecular diversity compared to elite bread wheat

The importance of wheat as a food crop makes it a major target for agricultural improvements. As one of the most widely grown cereal grains, together with maize and rice, wheat is the leading provider of calories in the global diet, constituting 29% of global cereal production in 2015. In the last few decades, however, yields have plateaued, suggesting that the green revolution, at least for wheat, might have run its course and that new sources of genetic variation are urgently required. The overall aim of our work was to identify novel variation that may then be used to enable the breeding process. As landraces are a potential source of such diversity, here we have characterized the A.E. Watkins Collection alongside a collection of elite accessions using two complementary high‐density and high‐throughput genotyping platforms. While our results show the importance of using the appropriate SNP collection to compare diverse accessions, they also show that the Watkins Collection contains a substantial amount of novel genetic diversity which has either not been captured in current breeding programmes or which has been lost through previous selection pressures. As a consequence of our analysis, we have identified a number of accessions which carry an array of novel alleles along with a number of interesting chromosome rearrangements which confirm the variable nature of the wheat genome.     

Genomic dissection of drought resistance in durum wheat × wild emmer wheat recombinant inbreed line population

Drought is the major factor limiting wheat productivity worldwide. The gene pool of wild emmer wheat, Triticum turgidum ssp. dicoccoides , harbours a rich allelic repertoire for morpho-physiological traits conferring drought resistance. The genetic and physiological bases of drought responses were studied here in a tetraploid wheat population of 152 recombinant inbreed lines (RILs), derived from a cross between durum wheat (cv. Langdon) and wild emmer (acc# G18-16), under contrasting water availabilities. Wide genetic variation was found among RILs for all studied traits. A total of 110 quantitative trait loci (QTLs) were mapped for 11 traits, with LOD score range of 3.0–35.4. Several QTLs showed environmental specificity, accounting for productivity and related traits under water-limited (20 QTLs) or well-watered conditions (15 QTLs), and in terms of drought susceptibility index (22 QTLs). Major genomic regions controlling productivity and related traits were identified on chromosomes 2B, 4A, 5A and 7B. QTLs for productivity were associated with QTLs for drought-adaptive traits, suggesting the involvement of several strategies in wheat adaptation to drought stress. Fifteen pairs of QTLs for the same trait were mapped to seemingly homoeologous positions, reflecting synteny between the A and B genomes. The identified QTLs may facilitate the use of wild alleles for improvement of drought resistance in elite wheat cultivars.     

A unified DNA sequence and non‐DNA sequence mapping model of complex traits

Increasing evidence shows that quantitative inheritance is based on both DNA sequence and non‐DNA sequence variants. However, how to simultaneously detect these variants from a mapping study has been unexplored, hampering our effort to illustrate the detailed genetic architecture of complex traits. We address this issue by developing a unified model of quantitative trait locus (QTL) mapping based on an open‐pollinated design composed of randomly sampling maternal plants from a natural population and their half‐sib seeds. This design forms a two‐level hierarchical platform for a joint linkage‐linkage disequilibrium analysis of population structure. The EM algorithm was implemented to estimate and test DNA sequence‐based effects and non‐DNA sequence‐based effects of QTLs. We applied this model to analyze genetic mapping data from the OP design of a gymnosperm coniferous species, Torreya grandis, identifying 25 significant DNA sequence and non‐DNA sequence QTLs for seedling height and diameter growth in different years. Results from computer simulation show that the unified model has good statistical properties and is powerful for QTL detection. Our model enables the tests of how a complex trait is affected differently by DNA‐based effects and non‐DNA sequence‐based transgenerational effects, thus allowing a more comprehensive picture of genetic architecture to be charted and quantified.     

Quantitative trait loci for growth and body size in the nine‐spined stickleback Pungitius pungitius L.

Body size is an ecologically important trait shown to be genetically variable both within and among different animal populations as revealed by quantitative genetic studies. However, few studies have looked into underlying genetic architecture of body size variability in the wild using genetic mapping methods. With the aid of quantitative trait loci (QTL) analyses based on 226 microsatellite markers, we mapped body size and growth rate traits in the nine‐spined stickleback (Pungitius pungitius) using an F₂‐intercross (n = 283 offspring) between size‐divergent populations. In total, 17 QTL locations were detected. The proportion of phenotypic variation explained by individual body size‐related QTL ranged from 3% to 12% and those related to growth parameters and increments from 3% to 10%. Several of the detected QTL affected either early or late growth. These results provide a solid starting point for more in depth investigations of structure and function of genomic regions involved in determination of body size in this popular model of ecological and evolutionary research.     

Genome‐wide association study for 13 agronomic traits reveals distribution of superior alleles in bread wheat from the Yellow and Huai Valley of China

Bread wheat is a leading cereal crop worldwide. Limited amount of superior allele loci restricted the progress of molecular improvement in wheat breeding. Here, we revealed new allelic variation distribution for 13 yield‐related traits in series of genome‐wide association studies (GWAS) using the wheat 90K genotyping assay, characterized in 163 bread wheat cultivars. Agronomic traits were investigated in 14 environments at three locations over 3 years. After filtering SNP data sets, GWAS using 20 689 high‐quality SNPs associated 1769 significant loci that explained, on average, ~20% of the phenotypic variation, both detected already reported loci and new promising genomic regions. Of these, repetitive and pleiotropic SNPs on chromosomes 6AS, 6AL, 6BS, 5BL and 7AS were significantly linked to thousand kernel weight, for example BS00021705_51 on 6BS and wsnp_Ex_c32624_41252144 on 6AS, with phenotypic variation explained (PVE) of ~24%, consistently identified in 12 and 13 of the 14 environments, respectively. Kernel length‐related SNPs were mainly identified on chromosomes 7BS, 6AS, 5AL and 5BL. Plant height‐related SNPs on chromosomes 4DS, 6DL, 2DS and 1BL were, respectively, identified in more than 11 environments, with averaged PVE of ~55%. Four SNPs were confirmed to be important genetic loci in two RIL populations. Based on repetivity and PVE, a total of 41 SNP loci possibly played the key role in modulating yield‐related traits of the cultivars surveyed. Distribution of superior alleles at the 41 SNP loci indicated that superior alleles were getting popular with time and modern cultivars had integrated many superior alleles, especially for peduncle length‐ and plant height‐related superior alleles. However, there were still 19 SNP loci showing less than percentages of 50% in modern cultivars, suggesting they should be paid more attention to improve yield‐related traits of cultivars in the Yellow and Huai wheat region. This study could provide useful information for dissection of yield‐related traits and valuable genetic loci for marker‐assisted selection in Chinese wheat breeding programme.     

Wheat tandem kinases provide insights on disease‐resistance gene flow and host–parasite co‐evolution

Stripe (yellow) rust, caused by the fungus Puccinia striiformis f. sp. tritici (Pst), is a destructive disease of wheat spread globally. Wild emmer wheat (Triticum turgidum ssp. dicoccoides; WEW) is known as a source for novel Pst resistance genes (R‐gene), but our knowledge on wheat‐Pst co‐evolution in natural populations is limited. Yr15 is a WEW (accession G25) gene, which confers a broad‐spectrum resistance to Pst, and encodes a tandem kinase‐pseudokinase protein designated as WTK1. Exon–intron comparisons of multiple WTK1 homoeologous and paralogous copies scattered in allopolyploid wheat genomes enabled us to develop functional molecular markers (FMMs), which were used for population genetic study. The functional allele (Wtk1) was absent in a worldwide collection of 513 wheat cultivars, except for 32 introgression lines with Yr15 from G25, as well as in 84% of the 382 tested WEW accessions collected across the Fertile Crescent. Yr15 was found to be distributed along a narrow axis from Mt Carmel to the Anti‐Lebanon Mountains ridge, mostly at elevations above c. 500 m, where the climatic conditions are favorable for disease development, therefore providing insights on gene flow and host–parasite co‐evolution in WEW natural habitats. Moreover, the worldwide absence of Wtk1 in cultivated wheat and in WEW natural populations from southeast Turkey, where wheat is believed to have been domesticated, proposes that Yr15 was rather left behind, than lost during domestication. Our results highlight the importance of conservation of WEW populations in their natural habitats for discovery of novel R‐genes and studies of host–parasite co‐evolution.     

Joint‐linkage mapping and GWAS reveal extensive genetic loci that regulate male inflorescence size in maize

Both insufficient and excessive male inflorescence size leads to a reduction in maize yield. Knowledge of the genetic architecture of male inflorescence is essential to achieve the optimum inflorescence size for maize breeding. In this study, we used approximately eight thousand inbreds, including both linkage populations and association populations, to dissect the genetic architecture of male inflorescence. The linkage populations include 25 families developed in the U.S. and 11 families developed in China. Each family contains approximately 200 recombinant inbred lines (RILs). The association populations include approximately 1000 diverse lines from the U.S. and China. All inbreds were genotyped by either sequencing or microarray. Inflorescence size was measured as the tassel primary branch number (TBN) and tassel length (TL). A total of 125 quantitative trait loci (QTLs) were identified (63 for TBN, 62 for TL) through linkage analyses. In addition, 965 quantitative trait nucleotides (QTNs) were identified through genomewide study (GWAS) at a bootstrap posterior probability (BPP) above a 5% threshold. These QTLs/QTNs include 24 known genes that were cloned using mutants, for example Ramosa3 (ra3), Thick tassel dwarf1 (td1), tasselseed2 (ts2), liguleless2 (lg2), ramosa1 (ra1), barren stalk1 (ba1), branch silkless1 (bd1) and tasselseed6 (ts6). The newly identified genes encode a zinc transporter (e.g. GRMZM5G838098 and GRMZM2G047762), the adapt in terminal region protein (e.g. GRMZM5G885628), O‐methyl‐transferase (e.g. GRMZM2G147491), helix‐loop‐helix (HLH) DNA‐binding proteins (e.g. GRMZM2G414252 and GRMZM2G042895) and an SBP‐box protein (e.g. GRMZM2G058588). These results provide extensive genetic information to dissect the genetic architecture of inflorescence size for the improvement of maize yield.     

Genome‐wide association mapping for female fertility traits in Danish and Swedish Holstein cattle

A genome‐wide association study was conducted using a mixed model analysis for QTL for fertility traits in Danish and Swedish Holstein cattle. The analysis incorporated 2,531 progeny tested bulls, and a total of 36 387 SNP markers on 29 bovine autosomes were used. Eleven fertility traits were analyzed for SNP association. Furthermore, mixed model analysis was used for association analyses where a polygenic effect was fitted as a random effect, and genotypes at single SNPs were successively included as a fixed effect in the model. The Bonferroni correction for multiple testing was applied to adjust the significance threshold. Seventy‐four SNP‐trait combinations showed chromosome‐wide significance, and five of these were significant genome‐wide. Twenty‐four QTL regions on 14 chromosomes were detected. Strong evidence for the presence of QTL that affect fertility traits were observed on chromosomes 3, 5, 10, 13, 19, 20, and 24. The QTL intervals were generally smaller than those described in earlier linkage studies. The identification of fertility trait‐associated SNPs and mapping of the corresponding QTL in small chromosomal regions reported here will facilitate searches for candidate genes and candidate polymorphisms.     

A composite method for mapping quantitative trait loci without interference of female achiasmatic and gender effects in silkworm,Bombyx mori

The silkworm, Bombyx mori, is an economically important insect that was domesticated more than 5000 years ago. Its major economic traits focused on by breeders are quantitative traits, and an accurate and efficient QTL mapping method is necessary to explore their genetic architecture. However, current widely used QTL mapping models are not well suited for silkworm because they ignore female achiasmate and gender effects. In this study, we propose a composite method combining rational population selection and special mapping methods to map QTL in silkworm. By determining QTL for cocoon shell weight (CSW), we demonstrated the effectiveness of this method. In the CSW mapping process, only 56 markers were used and five loci or chromosomes were detected, more than in previous studies. Additionally, loci on chromosomes 1 and 11 dominated and accounted for 35.10% and 15.03% of the phenotypic variance respectively. Unlike previous studies, epistasis was detected between loci on chromosomes 11 and 22. These mapping results demonstrate the power and convenience of this method for QTL mapping in silkworm, and this method may inspire the development of similar approaches for other species with special genetic characteristics.     

Genetic assignment of recruits reveals short‐ and long‐distance larval dispersal in Pocillopora damicornis on the Great Barrier Reef

Understanding connectivity of coral populations among and within reefs over ecologically significant timescales is essential for developing evidence‐based management strategies, including the design of marineprotected areas. Here, we present the first assessment of contemporary connectivity among populations of two Molecular Operational Taxonomic Units (MOTUs) of the brooding coral Pocillopora damicornis. We used individual‐based genetic assignment methods to identify the proportions of philopatric and migrant larval recruits, settling over 12 months at sites around Lizard Island (northern Great Barrier Reef [GBR]) and over 24 months at sites around the Palms Islands (central GBR). Overall, we found spatially and temporally variable rates of self‐recruitment and dispersal, demonstrating the importance of variation in local physical characteristics in driving dispersal processes. Recruitment patterns and inferred dispersal distances differed between the two P. damicornis MOTUs, with type α recruits exhibiting predominantly philopatric recruitment, while the majority of type β recruits were either migrants from identified putative source populations or assumed migrants based on genetic exclusion from all known populations. While P. damicornis invests much energy into brooding clonal larvae, we found that only 15% and 7% of type α and type β recruits, respectively, were clones of sampled adult colonies or other recruits, challenging the hypothesis that reproduction is predominantly asexual in this species on the GBR. We explain high rates of self‐recruitment and low rates of clonality in these MOTUs by suggesting that locally retained larvae originate predominantly from spawned gametes, while brooded larvae are mainly vagabonds.     

Genetic mapping shows intraspecific variation and transgressive segregation for caterpillar‐induced aphid resistance in maize

Plants in nature have inducible defences that sometimes lead to targeted resistance against particular herbivores, but susceptibility to others. The metabolic diversity and genetic resources available for maize (Zea mays) make this a suitable system for a mechanistic study of within‐species variation in such plant‐mediated interactions between herbivores. Beet armyworms (Spodoptera exigua) and corn leaf aphids (Rhopalosiphum maidis) are two naturally occurring maize herbivores with different feeding habits. Whereas chewing herbivore‐induced methylation of 2,4‐dihydroxy‐7‐methoxy‐1,4‐benzoxazin‐3‐one glucoside (DIMBOA‐Glc) to form 2‐hydroxy‐4,7‐dimethoxy‐1,4‐benzoxazin‐3‐one glucoside (HDMBOA‐Glc) promotes caterpillar resistance, lower DIMBOA‐Glc levels favour aphid reproduction. Thus, caterpillar‐induced DIMBOA‐Glc methyltransferase activity in maize is predicted to promote aphid growth. To test this hypothesis, the impact of S. exigua feeding on R. maidis progeny production was assessed using seventeen genetically diverse maize inbred lines. Whereas aphid progeny production was increased by prior caterpillar feeding on lines B73, Ki11, Ki3 and Tx303, it decreased on lines Ky21, CML103, Mo18W and W22. Genetic mapping of this trait in a population of B73 × Ky21 recombinant inbred lines identified significant quantitative trait loci on maize chromosomes 1, 7 and 10. There is a transgressive segregation for aphid resistance, with the Ky21 alleles on chromosomes 1 and 7 and the B73 allele on chromosome 10 increasing aphid progeny production. The chromosome 1 QTL coincides with a cluster of three maize genes encoding benzoxazinoid O‐methyltransferases that convert DIMBOA‐Glc to HDMBOA‐Glc. Gene expression studies and benzoxazinoid measurements indicate that S. exigua ‐induced responses in this pathway differentially affect R. maidis resistance in B73 and Ky21.     

Whole‐genome sequencing reveals absence of recent gene flow and separate demographic histories for Anopheles punctulatus mosquitoes in Papua New Guinea

Anopheles mosquitoes are the vectors of several human diseases including malaria. In many malaria endemic areas, several species of Anopheles coexist, sometimes in the form of related sibling species that are morphologically indistinguishable. Determining the size and organization of Anopheles populations, and possible ongoing gene flow among them is important for malaria control and, in particular, for monitoring the spread of insecticide resistance alleles. However, these parameters have been difficult to evaluate in most Anopheles species due to the paucity of genetic data available. Here, we assess the extent of contemporary gene flow and historical variations in population size by sequencing and de novo assembling the genomes of wild‐caught mosquitoes from four species of the Anopheles punctulatus group of Papua New Guinea. Our analysis of more than 50 Mb of orthologous DNA sequences revealed no evidence of contemporary gene flow among these mosquitoes. In addition, investigation of the demography of two of the An. punctulatus species revealed distinct population histories. Overall, our analyses suggest that, despite their similarities in morphology, behaviour and ecology, contemporary sympatric populations of An. punctulatus are evolving independently.     

The Inhibitor of wax 1 locus (Iw1) prevents formation of β‐ and OH‐β‐diketones in wheat cuticular waxes and maps to a sub‐cM interval on chromosome arm 2BS

Glaucousness is described as the scattering effect of visible light from wax deposited on the cuticle of plant aerial organs. In wheat, two dominant genes lead to non‐glaucous phenotypes: Inhibitor of wax 1 (Iw1) and Iw2. The molecular mechanisms and the exact extent (beyond visual assessment) by which these genes affect the composition and quantity of cuticular wax is unclear. To describe the Iw1 locus we used a genetic approach with detailed biochemical characterization of wax compounds. Using synteny and a large number of F₂ gametes, Iw1 was fine‐mapped to a sub‐cM genetic interval on wheat chromosome arm 2BS, which includes a single collinear gene from the corresponding Brachypodium and rice physical maps. The major components of flag leaf and peduncle cuticular waxes included primary alcohols, β‐diketones and n‐alkanes. Small amounts of C19–C27 alkyl and methylalkylresorcinols that have not previously been described in wheat waxes were identified. Using six pairs of BC₂F₃ near‐isogenic lines, we show that Iw1 inhibits the formation of β‐ and hydroxy‐β‐diketones in the peduncle and flag leaf blade cuticles. This inhibitory effect is independent of genetic background or tissue, and is accompanied by minor but consistent increases in n‐alkanes and C₂₄ primary alcohols. No differences were found in cuticle thickness and carbon isotope discrimination in near‐isogenic lines differing at Iw1.     

Development and characterization of EST‐SSR markers for the genus Rhododendron section Brachycalyx (Ericaceae)

Simple sequence repeat (SSR) markers were developed from expressed sequence tags (ESTs) for Rhododendron section Brachycalyx in order to elucidate its evolutionary processes and reproductive ecology. Nineteen polymorphic EST‐SSR markers were developed from EST libraries of R. amagianum and R. hyugaense. Polymorphisms for these markers were assessed using four species of section Brachycalyx. The number of alleles ranged from 1 to 14, and the observed and expected heterozygosity ranged from 0.000 to 0.931 and 0.000 to 0.904, respectively. The EST‐SSR markers developed in this study will be useful for elucidating population genetic structure and breeding systems in section Brachycalyx.     

A novel approach to parasite population genetics: Experimental infection reveals geographic differentiation,recombination and host‐mediated population structure in Pasteuria ramosa,a bacterial parasite of Daphnia

The population structure of parasites is central to the ecology and evolution of host‐parasite systems. Here, we investigate the population genetics of Pasteuria ramosa, a bacterial parasite of Daphnia. We used natural P. ramosa spore banks from the sediments of two geographically well‐separated ponds to experimentally infect a panel of Daphnia magna host clones whose resistance phenotypes were previously known. In this way, we were able to assess the population structure of P. ramosa based on geography, host resistance phenotype and host genotype. Overall, genetic diversity of P. ramosa was high, and nearly all infected D. magna hosted more than one parasite haplotype. On the basis of the observation of recombinant haplotypes and relatively low levels of linkage disequilibrium, we conclude that P. ramosa engages in substantial recombination. Isolates were strongly differentiated by pond, indicating that gene flow is spatially restricted. Pasteuria ramosa isolates within one pond were segregated completely based on the resistance phenotype of the host—a result that, to our knowledge, has not been previously reported for a nonhuman parasite. To assess the comparability of experimental infections with natural P. ramosa isolates, we examined the population structure of naturally infected D. magna native to one of the two source ponds. We found that experimental and natural infections of the same host resistance phenotype from the same source pond were indistinguishable, indicating that experimental infections provide a means to representatively sample the diversity of P. ramosa while reducing the sampling bias often associated with studies of parasite epidemics. These results expand our knowledge of this model parasite, provide important context for the large existing body of research on this system and will guide the design of future studies of this host‐parasite system.     

Developmental and genetic effects on behavioral and life‐history traits in a field cricket

A fundamental goal of evolutionary ecology is to identify the sources underlying trait variation on which selection can act. Phenotypic variation will be determined by both genetic and environmental factors, and adaptive phenotypic plasticity is expected when organisms can adjust their phenotypes to match environmental cues. Much recent research interest has focused on the relative importance of environmental and genetic factors on the expression of behavioral traits, in particular, and how they compare with morphological and life‐history traits. Little research to date examines the effect of development on the expression of heritable variation in behavioral traits, such as boldness and activity. We tested for genotype, environment, and genotype‐by‐environment differences in body mass, development time, boldness, and activity, using developmental density treatments combined with a quantitative genetic design in the sand field cricket (Gryllus firmus). Similar to results from previous work, animals reared at high densities were generally smaller and took longer to mature, and body mass and development time were moderately heritable. In contrast, neither boldness nor activity responded to density treatments, and they were not heritable. The only trait that showed significant genotype‐by‐environment differences was development time. It is possible that adaptive behavioral plasticity is not evident in this species because of the highly variable social environments it naturally experiences. Our results illustrate the importance of validating the assumption that behavioral phenotype reflects genetic patterns and suggest questions about the role of environmental instability in trait variation and heritability.     

Development and evaluation of a genome‐wide Coffee 8.5K SNP array and its application for high‐density genetic mapping and for investigating the origin of Coffea arabica L.

Coffee species such as Coffea canephora P. (Robusta) and C. arabica L. (Arabica) are important cash crops in tropical regions around the world. C. arabica is an allotetraploid (2n = 4x = 44) originating from a hybridization event of the two diploid species C. canephora and C. eugenioides (2n = 2x = 22). Interestingly, these progenitor species harbour a greater level of genetic variability and are an important source of genes to broaden the narrow Arabica genetic base. Here, we describe the development, evaluation and use of a single‐nucleotide polymorphism (SNP) array for coffee trees. A total of 8580 unique and informative SNPs were selected from C. canephora and C. arabica sequencing data, with 40% of the SNP located in annotated genes. In particular, this array contains 227 markers associated to 149 genes and traits of agronomic importance. Among these, 7065 SNPs (~82.3%) were scorable and evenly distributed over the genome with a mean distance of 54.4 Kb between markers. With this array, we improved the Robusta high‐density genetic map by adding 1307 SNP markers, whereas 945 SNPs were found segregating in the Arabica mapping progeny. A panel of C. canephora accessions was successfully discriminated and over 70% of the SNP markers were transferable across the three species. Furthermore, the canephora‐derived subgenome of C. arabica was shown to be more closely related to C. canephora accessions from northern Uganda than to other current populations. These validated SNP markers and high‐density genetic maps will be useful to molecular genetics and for innovative approaches in coffee breeding.