Limited haplotype diversity underlies polygenic trait architecture across 70 years of wheat breeding.

BACKGROUND: Selection has dramatically shaped genetic and phenotypic variation in bread wheat. We can assess the genomic basis of historical phenotypic changes, and the potential for future improvement, using experimental populations that attempt to undo selection through the randomizing effects of recombination. RESULTS: We bred the NIAB Diverse MAGIC multi-parent population comprising over 500 recombinant inbred lines, descended from sixteen historical UK bread wheat varieties released between 1935 and 2004. We sequence the founders' genes and promoters by capture, and the MAGIC population by low-coverage whole-genome sequencing. We impute 1.1 M high-quality SNPs that are over 99% concordant with array genotypes. Imputation accuracy only marginally improves when including the founders' genomes as a haplotype reference panel. Despite capturing 73% of global wheat genetic polymorphism, 83% of genes cluster into no more than three haplotypes. We phenotype 47 agronomic traits over 2 years and map 136 genome-wide significant associations, concentrated at 42 genetic loci with large and often pleiotropic effects. Around half of these overlap known quantitative trait loci. Most traits exhibit extensive polygenicity, as revealed by multi-locus shrinkage modelling. CONCLUSIONS: Our results are consistent with a gene pool of low haplotypic diversity, containing few novel loci of large effect. Most past, and projected future, phenotypic changes arising from existing variation involve fine-scale shuffling of a few haplotypes to recombine dozens of polygenic alleles of small effect. Moreover, extensive pleiotropy means selection on one trait will have unintended consequences, exemplified by the negative trade-off between yield and protein content, unless selection and recombination can break unfavorable trait-trait associations.

A high-throughput delayed fluorescence method reveals underlying differences in the control of circadian rhythms in Triticum aestivum and Brassica napus.

BACKGROUND: A robust circadian clock has been implicated in plant resilience, resource-use efficiency, competitive growth and yield. A huge number of physiological processes are under circadian control in plants including: responses to biotic and abiotic stresses; flowering time; plant metabolism; and mineral uptake. Understanding how the clock functions in crops such as Triticum aestivum (bread wheat) and Brassica napus (oilseed rape) therefore has great agricultural potential. Delayed fluorescence (DF) imaging has been shown to be applicable to a wide range of plant species and requires no genetic transformation. Although DF has been used to measure period length of both mutants and wild ecotypes of Arabidopsis, this assay has never been systematically optimised for crop plants. The physical size of both B. napus and T. aestivum led us to develop a representative sampling strategy which enables high-throughput imaging of these crops. RESULTS: In this study, we describe the plant-specific optimisation of DF imaging to obtain reliable circadian phenotypes with the robustness and reproducibility to detect diverging periods between cultivars of the same species. We find that the age of plant material, light regime and temperature conditions all significantly effect DF rhythms and describe the optimal conditions for measuring robust rhythms in each species. We also show that sections of leaf can be used to obtain period estimates with improved throughput for larger sample size experiments. CONCLUSIONS: We present an optimized protocol for high-throughput phenotyping of circadian period specific to two economically valuable crop plants. Application of this method revealed significant differences between the periods of several widely grown elite cultivars. This method also identified intriguing differential responses of circadian rhythms in T. aestivum compared to B. napus; specifically the dramatic change to rhythm robustness when plants were imaged under constant light versus constant darkness. This points towards diverging networks underlying circadian control in these two species.

Analysis of the recombination landscape of hexaploid bread wheat reveals genes controlling recombination and gene conversion frequency.

BACKGROUND: Sequence exchange between homologous chromosomes through crossing over and gene conversion is highly conserved among eukaryotes, contributing to genome stability and genetic diversity. A lack of recombination limits breeding efforts in crops; therefore, increasing recombination rates can reduce linkage drag and generate new genetic combinations. RESULTS: We use computational analysis of 13 recombinant inbred mapping populations to assess crossover and gene conversion frequency in the hexaploid genome of wheat (Triticum aestivum). We observe that high-frequency crossover sites are shared between populations and that closely related parents lead to populations with more similar crossover patterns. We demonstrate that gene conversion is more prevalent and covers more of the genome in wheat than in other plants, making it a critical process in the generation of new haplotypes, particularly in centromeric regions where crossovers are rare. We identify quantitative trait loci for altered gene conversion and crossover frequency and confirm functionality for a novel RecQ helicase gene that belongs to an ancient clade that is missing in some plant lineages including Arabidopsis. CONCLUSIONS: This is the first gene to be demonstrated to be involved in gene conversion in wheat. Harnessing the RecQ helicase has the potential to break linkage drag utilizing widespread gene conversions.

Integrating genomic resources to present full gene and putative promoter capture probe sets for bread wheat.

BACKGROUND: Whole-genome shotgun resequencing of wheat is expensive because of its large, repetitive genome. Moreover, sequence data can fail to map uniquely to the reference genome, making it difficult to unambiguously assign variation. Resequencing using target capture enables sequencing of large numbers of individuals at high coverage to reliably identify variants associated with important agronomic traits. Previous studies have implemented complementary DNA/exon or gene-based probe sets in which the promoter and intron sequence is largely missing alongside newly characterized genes from the recent improved reference sequences. RESULTS: We present and validate 2 gold standard capture probe sets for hexaploid bread wheat, a gene and a putative promoter capture, which are designed using recently developed genome sequence and annotation resources. The captures can be combined or used independently. We demonstrate that the capture probe sets effectively enrich the high-confidence genes and putative promoter regions that were identified in the genome alongside a large proportion of the low-confidence genes and associated promoters. Finally, we demonstrate successful sample multiplexing that allows generation of adequate sequence coverage for single-nucleotide polymorphism calling while significantly reducing cost per sample for gene and putative promoter capture. CONCLUSIONS: We show that a capture design employing an "island strategy" can enable analysis of the large gene/putative promoter space of wheat with only 2 × 160 Mbp probe sets. Furthermore, these assays extend the regions of the wheat genome that are amenable to analyses beyond its exome, providing tools for detailed characterization of these regulatory regions in large populations.

Harnessing genetic potential of wheat germplasm banks through impact-oriented-prebreeding for future food and nutritional security.

The value of exotic wheat genetic resources for accelerating grain yield gains is largely unproven and unrealized. We used next-generation sequencing, together with multi-environment phenotyping, to study the contribution of exotic genomes to 984 three-way-cross-derived (exotic/elite1//elite2) pre-breeding lines (PBLs). Genomic characterization of these lines with haplotype map-based and SNP marker approaches revealed exotic specific imprints of 16.1 to 25.1%, which compares to theoretical expectation of 25%. A rare and favorable haplotype (GT) with 0.4% frequency in gene bank identified on chromosome 6D minimized grain yield (GY) loss under heat stress without GY penalty under irrigated conditions. More specifically, the 'T' allele of the haplotype GT originated in Aegilops tauschii and was absent in all elite lines used in study. In silico analysis of the SNP showed hits with a candidate gene coding for isoflavone reductase IRL-like protein in Ae. tauschii. Rare haplotypes were also identified on chromosomes 1A, 6A and 2B effective against abiotic/biotic stresses. Results demonstrate positive contributions of exotic germplasm to PBLs derived from crosses of exotics with CIMMYT's best elite lines. This is a major impact-oriented pre-breeding effort at CIMMYT, resulting in large-scale development of PBLs for deployment in breeding programs addressing food security under climate change scenarios.

Prospects for the use of biological control agents against Anoplophora in Europe.

This review summarises the literature on the biological control of Anoplophora spp. (Coleoptera: Cerambycidae) and discusses its potential for use in Europe. Entomopathogenic fungi: Beauveria brongniartii Petch (Hypocreales: Cordycipitaceae) has already been developed into a commercial product in Japan, and fungal infection results in high mortality rates. Parasitic nematodes: Steinernema feltiae Filipjev (Rhabditida: Steinernematidae) and Steinernema carpocapsae Weiser have potential for use as biopesticides as an alternative to chemical treatments. Parasitoids: a parasitoid of Anoplophora chinensis Forster, Aprostocetus anoplophorae Delvare (Hymenoptera: Eulophidae), was discovered in Italy in 2002 and has been shown to be capable of parasitising up to 72% of A. chinensis eggs; some native European parasitoid species (e.g. Spathius erythrocephalus) also have potential to be used as biological control agents. Predators: two woodpecker (Piciformis: Picidae) species that are native to Europe, Dendrocopos major Beicki and Picus canus Gmelin, have been shown to be effective at controlling Anoplophora glabripennis Motschulsky in Chinese forests. The removal and destruction of infested and potentially infested trees is the main eradication strategy for Anoplophora spp. in Europe, but biological control agents could be used in the future to complement other management strategies, especially in locations where eradication is no longer possible.

The stochastic silencing phenotype of Arabidopsis morc6 mutants reveals a role in efficient RNA-directed DNA methylation.

The RNA-directed DNA methylation (RdDM) pathway is of central importance to the initiation and maintenance of transcriptional gene silencing in plants. DNA methylation is directed to target sequences by a mechanism that involves production of small RNAs by RNA polymerase IV and long non-coding RNAs by RNA polymerase V. DNA methylation then leads to recruitment of histone-modifying enzymes, followed by establishment of a silenced chromatin state. Recently MORC6, a member of the microrchidia (MORC) family of adenosine triphosphatases (ATPases), has been shown to be involved in transcriptional gene silencing. However, reports differ regarding whether MORC6 is involved in RdDM itself or acts downstream of DNA methylation to enable formation of higher-order chromatin structure. Here we demonstrate that MORC6 is required for efficient RdDM at some target loci, and, using a GFP reporter system, we found that morc6 mutants show a stochastic silencing phenotype. By using cell sorting to separate silenced and unsilenced cells, we show that release of silencing at this locus is associated with a loss of DNA methylation. Thus our data support a view that MORC6 influences RdDM and that it is not acting downstream of DNA methylation. For some loci, efficient initiation or maintenance of DNA methylation may depend on the ability to form higher-order chromatin structure.