Inter-annual and spatial climatic variability have led to a balance between local fluctuating selection and wide-range directional selection in a perennial grass species.

BACKGROUND AND AIMS: The persistence of a plant population under a specific local climatic regime requires phenotypic adaptation with underlying particular combinations of alleles at adaptive loci. The level of allele diversity at adaptive loci within a natural plant population conditions its potential to evolve, notably towards adaptation to a change in climate. Investigating the environmental factors that contribute to the maintenance of adaptive diversity in populations is thus worthwhile. Within-population allele diversity at adaptive loci can be partly driven by the mean climate at the population site but also by its temporal variability. METHODS: The effects of climate temporal mean and variability on within-population allele diversity at putatively adaptive quantitative trait loci (QTLs) were evaluated using 385 natural populations of Lolium perenne (perennial ryegrass) collected right across Europe. For seven adaptive traits related to reproductive phenology and vegetative potential growth seasonality, the average within-population allele diversity at major QTLs (HeA) was computed. KEY RESULTS: Significant relationships were found between HeA of these traits and the temporal mean and variability of the local climate. These relationships were consistent with functional ecology theory. CONCLUSIONS: Results indicated that temporal variability of local climate has likely led to fluctuating directional selection, which has contributed to the maintenance of allele diversity at adaptive loci and thus potential for further adaptation.

Integrated analysis of yield response and early stage biochemical, molecular, and gene expression profiles of pre-breeding rice lines under water deficit stress.

Breeding high yielding water-deficit tolerant rice is considered a primary goal for achieving the objectives of the sustainable development goals, 2030. However, evaluating the performance of the pre-breeding-promising parental-lines for water deficit tolerance prior to their incorporation in the breeding program is crucial for the success of the breeding programs. The aim of the current investigation is to assess the performance of a set of pre-breeding lines compared with their parents. To achieve this goal a set of 7 pre-breeding rice lines along with their parents (5 genotypes) were field evaluated under well-irrigated and water-stress conditions. Water stress was applied by flush irrigation every 12 days without keeping standing water after irrigation. Based on the field evaluation results, a pre-breeding line was selected to conduct physiological and expression analysis of drought related genes at the green house. Furthermore, a greenhouse trial was conducted in pots, where the genotypes were grown under well and stress irrigation conditions at seedling stage for physiological analysis and expression profiling of the genotypes. Results indicated that the pre-breeding lines which were high yielding under water shortage stress showed low drought susceptibility index. Those lines exhibited high proline, SOD, TSS content along with low levels of MDA content in their leaves. Moreover, the genotypes grain yield positively correlated with proline, SOD, TSS content in their leaves. The SSR markers RM22, RM525, RM324 and RM3805 were able to discriminate the tolerant parents from the sensitive one. Expression levels of the tested drought responsive genes revealed the upregulation of OsLEA3, OsAPX2, OsNAC1, OSDREB2A, OsDREB1C, OsZIP23, OsP5CS, OsAHL1 and OsCATA genes in response to water deficit stress as compared to their expression under normal irrigated condition. Taken together among the tested pre-breeding lines the RBL112 pre-breeding line is high yielding under water-deficit and could be used as donor for high yielding genes in the breeding for water deficit resistance. This investigation withdraws attention to evaluate the promising pre-breeding lines before their incorporation in the water deficit stress breeding program.

Divergent evolution drives high diversity of toll-like receptors (TLRs) in passerine birds: Buntings and finches.

Toll-like receptors (TLRs) form a key component of animal innate immunity, being responsible for recognition of conserved microbial structures. As such, TLRs may be subject to diversifying and balancing selection, which maintains allelic variation both within and between populations. However, most research on TLRs in non-model avian species is focused on bottlenecked populations with depleted genetic variation. Here, we assessed variation at the extracellular domains of three TLR genes (TLR1LA, TLR3, TLR4) across eleven species from two passerine families of buntings (Emberizidae) and finches (Fringillidae), all having large breeding population sizes (millions of individuals). We found extraordinary TLR polymorphism in our study taxa, with >100 alleles detected at TLR1LA and TLR4 across species and high haplotype diversity (>0.75) in several species. Despite recent species divergence, no nucleotide allelic variants were shared between species, suggesting rapid TLR evolution. Higher variation at TLR1LA and TLR4 than TLR3 was associated with a stronger signal of diversifying selection, as measured with nucleotide substitutions rates and the number of positively selected sites (PSS). Structural protein modelling of TLRs showed that some PSS detected within TLR1LA and TLR4 were previously recognized as functionally important sites or were located in their proximity, possibly affecting ligand recognition. Furthermore, we identified PSS responsible for major surface electrostatic charge clustering, which may indicate their adaptive importance. Our study provides compelling evidence for the divergent evolution of TLR genes in buntings and finches and indicates that high TLR variation may be adaptively maintained via diversifying selection acting on functional ligand binding sites.

Phased, chromosome-scale genome assemblies of tetraploid potato reveal a complex genome, transcriptome, and predicted proteome landscape underpinning genetic diversity.

Cultivated potato is a clonally propagated autotetraploid species with a highly heterogeneous genome. Phased assemblies of six cultivars including two chromosome-scale phased genome assemblies revealed extensive allelic diversity, including altered coding and transcript sequences, preferential allele expression, and structural variation that collectively result in a highly complex transcriptome and predicted proteome, which are distributed across the homologous chromosomes. Wild species contribute to the extensive allelic diversity in tetraploid cultivars, demonstrating ancestral introgressions predating modern breeding efforts. As a clonally propagated autotetraploid that undergoes limited meiosis, dysfunctional and deleterious alleles are not purged in tetraploid potato. Nearly a quarter of the loci bore mutations are predicted to have a high negative impact on protein function, complicating breeder's efforts to reduce genetic load. The StCDF1 locus controls maturity, and analysis of six tetraploid genomes revealed that 12 allelic variants of StCDF1 are correlated with maturity in a dosage-dependent manner. Knowledge of the complexity of the tetraploid potato genome with its rampant structural variation and embedded deleterious and dysfunctional alleles will be key not only to implementing precision breeding of tetraploid cultivars but also to the construction of homozygous, diploid potato germplasm containing favorable alleles to capitalize on heterosis in F1 hybrids.

The Significance of RHD Genotyping and Characteristic Analysis in Chinese RhD Variant Individuals.

Background: RhD is the most important and complex blood group system because of its highly polymorphic and immunogenic nature. RhD variants can induce immune response by allogeneic transfusion, organ transplantation, and fetal immunity. The transfusion strategies are different for RhD variants formed by various alleles. Therefore, extensive investigation of the molecular mechanism underlying RhD variants is critical for preventing immune-related blood transfusion reactions and fetal immunity. Methods: RhD variants were collected from donors and patients in Zhejiang Province, China. The phenotypes were classified using the serologic method. The full coding regions of RHD gene were analyzed using the PCR-SBT method. The multiplex ligation-dependent probe amplification (MLPA) assay was used to analyze the genotype and gene copy number. SWISS-MODLE and PyMOL software were used to analyze 3D structures of RhD caused by the variant alleles. The effect of non-synonymous substitutions was predicted using Polymorphism Phenotyping algorithm (PolyPhen-2), Sorting Intolerant From Tolerant (SIFT), and Protein Variation Effect Analyzer (PROVEAN) software. Results: In the collected RhD variants, 28 distinct RHD variant alleles were identified, including three novel variant alleles. RH-MLPA assay is advantageous for determining the copy number of RHD gene. 3D homology modeling predicted that protein conformation was disrupted and may explain RhD epitope differential expression. A total of 14 non-synonymous mutations were determined to be detrimental to the protein structure. Discussion: We revealed the diversity of RHD alleles present in eastern Chinese RhD variants. The bioinformatics of these variant alleles extended our knowledge of RhD variants, which was crucial for evaluating their impact to guide transfusion support and avoid immune-related blood transfusion reactions.

Whole-Genome Resequencing Identifies KIT New Alleles That Affect Coat Color Phenotypes in Pigs.

The Duroc × (Landrace × Large White) hybrid pig (DLY) is the most popular commercial pig used in the Chinese pig industry. DLY pigs are usually white but sometimes show colored phenotypes. Colored DLY pigs are not favored by slaughterhouses and retailers, thus causing certain economic losses to farmers in China. In this study, we first conducted a genome-wide association study and RNA sequencing to demonstrate that KIT variants are responsible for diversifying coat color phenotypes segregating in a DLY population. We then defined the precise sizes and locations of four duplications (DUP1-4), four candidate causative mutations at the KIT locus, in the pig reference genome using the whole-genome sequence data of representative colored individuals. The sequence data also enabled us to identify a list of new KIT alleles. By investigating the association between these new alleles and coat color phenotypes, we provide further evidence that DUP2 is another causative mutation for the solid white coat color in pigs. DUP1 (the KIT gene duplication), DUP2 and the splice mutation are all required for the manifestation of a solid white coat color. DUP4 had a more significant effect on the formation of the belt phenotype compared with DUP3. Given the necessity of DUP2 for the solid white coat color, we detected IN /IN homozygotes lacking DUP2 in Large White and Landrace pigs and found that French Landrace pigs had the highest frequency (8.98%) of IN /IN individuals. This study not only advances our understanding of the molecular mechanism of the color phenotype in pigs, but also establishes a simple and accurate method for the screening of KIT IN /IN homozygotes in Large White and Landrace that would cause colored DLY pigs.

SNP Discovery and Genetic Variation of Candidate Genes Relevant to Heat Tolerance and Agronomic Traits in Natural Populations of Sand Rice (Agriophyllum squarrosum).

The extreme stress tolerance and high nutritional value of sand rice (Agriophyllum squarrosum) make it attractive for use as an alternative crop in response to concerns about ongoing climate change and future food security. However, a lack of genetic information hinders understanding of the mechanisms underpinning the morphological and physiological adaptations of sand rice. In the present study, we sequenced and analyzed the transcriptomes of two individuals representing semi-arid [Naiman (NM)] and arid [Shapotou (SPT)] sand rice genotypes. A total of 105,868 pairwise single nucleotide polymorphisms (SNPs) distributed in 24,712 Unigenes were identified among SPT and NM samples; the average SNP frequency was 0.3% (one SNP per 333 base pair). Characterization of gene annotation demonstrated that variations in genes involved in DNA recombination were associated with the survival of the NM population in the semi-arid environment. A set of genes predicted to be relevant to heat stress response and agronomic traits was functionally annotated using the accumulated knowledge from Arabidopsis and several crop plants, including rice, barley, maize, and sorghum. Four candidate genes related to heat tolerance (heat-shock transcription factor, HsfA1d), seed size (DA1-Related, DAR1), and flowering (early flowering 3, ELF3 and late elongated hypocotyl, LHY) were subjected to analysis of the genetic diversity in 10 natural populations, representing the core germplasm resource across the area of sand rice distribution in China. Only one SNP was detected in each of HsfA1d and DAR1, among 60 genotypes, with two in ELF3 and four in LHY. Nucleotide diversity ranged from 0.00032 to 0.00118. Haplotype analysis indicated that the NM population carried a specific allele for all four genes, suggesting that divergence has occurred between NM and other populations. These four genes could be further analyzed to determine whether they are associated with phenotype variation and identify alleles favorable for sand rice breeding.

Next generation breeding.

The genomic revolution of the past decade has greatly improved our understanding of the genetic make-up of living organisms. The sequencing of crop genomes has completely changed our vision and interpretation of genome organization and evolution. Re-sequencing allows the identification of an unlimited number of markers as well as the analysis of germplasm allelic diversity based on allele mining approaches. High throughput marker technologies coupled with advanced phenotyping platforms provide new opportunities for discovering marker-trait associations which can sustain genomic-assisted breeding. The availability of genome sequencing information is enabling genome editing (site-specific mutagenesis), to obtain gene sequences desired by breeders. This review illustrates how next generation sequencing-derived information can be used to tailor genomic tools for different breeders' needs to revolutionize crop improvement.

Population genetics of self-incompatibility in a clade of relict cliff-dwelling plant species.

The mating systems of species in small or fragmented populations impact upon their persistence. Small self-incompatible (SI) populations risk losing S allele diversity, responsible for the SI response, by drift thereby limiting mate availability and leading to population decline or SI system breakdown. But populations of relict and/or endemic species have resisted these demographic conditions over long periods suggesting their mating systems have adapted. To address a lack of empirical data on this topic, we studied the SI systems of three relict cliff-dwelling species of Sonchus section Pustulati (Asteraceae): S. masguindalii, S. fragilis and S. pustulatus in the western Mediterranean region. We performed controlled pollinations within and between individuals to measure index of SI (ISI) expression and identify S alleles in multiple population samples. Sonchus masguindalii and S. pustulatus showed strong SI (ISI = 0.6-1.0) compared to S. fragilis (ISI = 0.1-0.7). Just five S alleles were estimated for Spanish S. pustulatus and a moderate 11-15 S alleles for Moroccan S. pustulatus and S. fragilis, respectively. The fact that autonomous fruit set was generally improved by active self-pollination in self-compatible S. fragilis suggests that individuals with weak SI can show a wide range of outcrossing levels dependent on the degree of self or outcross pollen that pollinators bear. We conclude that frequent S allele dominance interactions that mask the incompatibility interactions of recessive S alleles leading to higher mate availability and partial breakdown of SI leading to mixed mating, both contribute to reproductive resilience in this group.