Genome-Wide Association Studies Using Single Nucleotide Polymorphism Markers Developed by Re-Sequencing of the Genomes of Cultivated Tomato

With the aim of understanding relationship between genetic and phenotypic variations in cultivated tomato, single nucleotide polymorphism (SNP) markers covering the whole genome of cultivated tomato were developed and genome-wide association studies (GWAS) were performed. The whole genomes of six tomato lines were sequenced with the ABI-5500xl SOLiD sequencer. Sequence reads covering ∼13.7× of the genome for each line were obtained, and mapped onto tomato reference genomes (SL2.40) to detect ∼1.5 million SNP candidates. Of the identified SNPs, 1.5% were considered to confer gene functions. In the subsequent Illumina GoldenGate assay for 1536 SNPs, 1293 SNPs were successfully genotyped, and 1248 showed polymorphisms among 663 tomato accessions. The whole-genome linkage disequilibrium (LD) analysis detected highly biased LD decays between euchromatic (58 kb) and heterochromatic regions (13.8 Mb). Subsequent GWAS identified SNPs that were significantly associated with agronomical traits, with SNP loci located near genes that were previously reported as candidates for these traits. This study demonstrates that attractive loci can be identified by performing GWAS with a large number of SNPs obtained from re-sequencing analysis.     

The Genetic Basis of Natural Variation in Caenorhabditis elegans Telomere Length

Telomeres are involved in the maintenance of chromosomes and the prevention of genome instability. Despite this central importance, significant variation in telomere length has been observed in a variety of organisms. The genetic determinants of telomere-length variation and their effects on organismal fitness are largely unexplored. Here, we describe natural variation in telomere length across the Caenorhabditis elegans species. We identify a large-effect variant that contributes to differences in telomere length. The variant alters the conserved oligonucleotide/oligosaccharide-binding fold of protection of telomeres 2 (POT-2), a homolog of a human telomere-capping shelterin complex subunit. Mutations within this domain likely reduce the ability of POT-2 to bind telomeric DNA, thereby increasing telomere length. We find that telomere-length variation does not correlate with offspring production or longevity in C. elegans wild isolates, suggesting that naturally long telomeres play a limited role in modifying fitness phenotypes in C. elegans.     

Quantitative Genetic Analysis Indicates Natural Selection on Leaf Phenotypes Across Wild Tomato Species (Solanum sect. Lycopersicon; Solanaceae)

Adaptive evolution requires both raw genetic material and an accessible path of high fitness from one fitness peak to another. In this study, we used an introgression line (IL) population to map quantitative trait loci (QTL) for leaf traits thought to be associated with adaptation to precipitation in wild tomatoes (Solanum sect. Lycopersicon; Solanaceae). A QTL sign test showed that several traits likely evolved under directional natural selection. Leaf traits correlated across species do not share a common genetic basis, consistent with a scenario in which selection maintains trait covariation unconstrained by pleiotropy or linkage disequilibrium. Two large effect QTL for stomatal distribution colocalized with key genes in the stomatal development pathway, suggesting promising candidates for the molecular bases of adaptation in these species. Furthermore, macroevolutionary transitions between vastly different stomatal distributions may not be constrained when such large-effect mutations are available. Finally, genetic correlations between stomatal traits measured in this study and data on carbon isotope discrimination from the same ILs support a functional hypothesis that the distribution of stomata affects the resistance to CO₂ diffusion inside the leaf, a trait implicated in climatic adaptation in wild tomatoes. Along with evidence from previous comparative and experimental studies, this analysis indicates that leaf traits are an important component of climatic niche adaptation in wild tomatoes and demonstrates that some trait transitions between species could have involved few, large-effect genetic changes, allowing rapid responses to new environmental conditions.     

Molecular Polymorphism and Morphometrics of Species of the Heterodera avenae Group in Syria and Turkey

Molecular characterization of the three most common cereal cyst nematode species of the Heterodera avenae group (H. avenae, H. filipjevi, and H. latipons), originating from various locations in major cereal-cultivating areas in Syria and Turkey, showed distinct restriction fragment patterns of the ITS-rDNA following PCR amplification and RFLP digestion with four endonucleases (Hae III, Hinf I, Ita I, and Pst I). Genetic dissimilarity within H. avenae group populations increased in comparison with H. avenae and other species; it was 0.164 with H. filipjevi and 0.354 with H. latipons populations. No intraspecific polymorphism was observed within H. latipons or H. filipjevi populations. Principal component analysis revealed contrasted correlations among 12 morphological parameters of cysts and juveniles of the three Heterodera species that separated them and distinguished differences within populations of H. latipons. Our results showed a clear separation of the three cyst nematode species on cereal using a conventional method for classification and molecular tests, and confirmed the congruence between genetics and morphological traits.     

Separation of Three Species of Ditylenchus and Some Host Races of D. dipsaci by Restriction Fragment Length Polymorphism

This study examined the ribosomal cistron of Ditylenchus destructor, D. myceliophagus and seven host races of D. dipsaci from different geographic locations. The three species showed restriction fragment length polymorphisms (RFLPs) in the ribosomal cistron, the 18S rDNA gene, and the ribosomal internal transcribed spacer (ITS). Southern blot analysis with a 7.5-kb ribosomal cistron probe differentiated the five host races of D. dipsaci examined. Polymerase chain reaction (PCR) amplification of the ITS, followed by digestion with some restriction endonucleases (but not others), produced restriction fragments diagnostic of the giant race. Because the PCR product from D. myceliophagus and the host races of D. dipsaci was about 900 base pairs and the ITS size in D. destructor populations was 1,200 base pairs, mixtures of populations could be detected by PCR amplification. ITS fragments differentiated between D. dipsaci and Aphelenchoides rhyntium in mixed populations. This study establishes the feasibility of differentiation of the host races of D. dipsaci by probing Southern blots with the whole ribosomal cistron.     

Molecular Polymorphism in the MTA and MTB Mating Type Genes of Tetrahymena thermophila and Related Asexual Species

Each of the seven mating types of Tetrahymena thermophila is determined by a pair of large genes, MTA and MTB, whose expression peaks at early conjugation. Each protein consists of a mating‐type specific domain and a common transmembrane domain. To assess variation in natural populations, regions of both domains from wild isolates expressing mating types V and VII were analyzed. Corresponding regions of amicronucleates incapable of mating also were examined. MTA and MTB showed high haplotype diversity, with greater sequence variation in MTB. Mating type VII was less variable than mating type V, suggesting more recent origin. No polymorphism distinguished between mat1‐ and mat2‐like alleles encoding different arrays of mating types, nor did polymorphisms give evidence of population structure. MTA and MTB variants have different phylogenies, suggesting independent rather than concerted evolution, and are under weak purifying selection. Codon usage is less biased than for housekeeping genes, and reassigned glutamine encoding stop codons are preferentially used. Amicronucleate T. thermophila and closely related nsp15 and nsp25 have higher levels of nucleotide and amino acid substitution, consistent with cox1 distances. The results suggest that complete sequencing of mating type genes of wild isolates coupled with functional analysis will be informative.     

The Effects of Background and Interference Selection on Patterns of Genetic Variation in Subdivided Populations

It is well known that most new mutations that affect fitness exert deleterious effects and that natural populations are often composed of subpopulations (demes) connected by gene flow. To gain a better understanding of the joint effects of purifying selection and population structure, we focus on a scenario where an ancestral population splits into multiple demes and study neutral diversity patterns in regions linked to selected sites. In the background selection regime of strong selection, we first derive analytic equations for pairwise coalescent times and F_ST as a function of time after the ancestral population splits into two demes and then construct a flexible coalescent simulator that can generate samples under complex models such as those involving multiple demes or nonconservative migration. We have carried out extensive forward simulations to show that the new methods can accurately predict diversity patterns both in the nonequilibrium phase following the split of the ancestral population and in the equilibrium between mutation, migration, drift, and selection. In the interference selection regime of many tightly linked selected sites, forward simulations provide evidence that neutral diversity patterns obtained from both the nonequilibrium and equilibrium phases may be virtually indistinguishable for models that have identical variance in fitness, but are nonetheless different with respect to the number of selected sites and the strength of purifying selection. This equivalence in neutral diversity patterns suggests that data collected from subdivided populations may have limited power for differentiating among the selective pressures to which closely linked selected sites are subject.     

Demographic History and Natural Selection Shape Patterns of Deleterious Mutation Load and Barriers to Introgression across Populus Genome

Hybridization and resulting introgression are important processes shaping the tree of life and appear to be far more common than previously thought. However, how the genome evolution was shaped by various genetic and evolutionary forces after hybridization remains unresolved. Here we used whole-genome resequencing data of 227 individuals from multiple widespread Populus species to characterize their contemporary patterns of hybridization and to quantify genomic signatures of past introgression. We observe a high frequency of contemporary hybridization and confirm that multiple previously ambiguous species are in fact F₁ hybrids. Seven species were identified, which experienced different demographic histories that resulted in strikingly varied efficacy of selection and burdens of deleterious mutations. Frequent past introgression has been found to be a pervasive feature throughout the speciation of these Populus species. The retained introgressed regions, more generally, tend to contain reduced genetic load and to be located in regions of high recombination. We also find that in pairs of species with substantial differences in effective population size, introgressed regions are inferred to have undergone selective sweeps at greater than expected frequencies in the species with lower effective population size, suggesting that introgression likely have higher potential to provide beneficial variation for species with small populations. Our results, therefore, illustrate that demography and recombination have interplayed with both positive and negative selection in determining the genomic evolution after hybridization.     

Polymorphic Foraging Behavior Among Caenorhabditis elegans: Frequency- and Density-Dependent Selection

Strains of Caenorhabditis elegans obtained from their natural soil environment exhibit one of two forms of foraging behavior. Some strains forage solitarily and disperse evenly on a bacterial lawn. Other strains move rapidly until they encounter groups of conspecifics, and then slow their movement and join the group. Strains expressing these behaviors are globally widespread and have been isolated from the same location, suggesting a foraging polymorphism. We hypothesized that density-dependent selection maintains both foraging alleles in populations. Alternatively, both foraging alleles could be retained in populations through frequency-dependent selection. We tested both of these hypotheses by manipulating strain density and frequency, and observing changes in population density over time. Our results indicated that neither density- nor frequency-dependent selection appears to be responsible for the observed polymorphism. The clumping strain consistently out-competed the solitary strain over all treatment levels. We suggest other potential factors that may maintain both alleles in populations.     

Exploiting the Extraordinary Genetic Polymorphism of Ciona for Developmental Genetics with Whole Genome Sequencing

Studies in tunicates such as Ciona have revealed new insights into the evolutionary origins of chordate development. Ciona populations are characterized by high levels of natural genetic variation, between 1 and 5%. This variation has provided abundant material for forward genetic studies. In the current study, we make use of deep sequencing and homozygosity mapping to map spontaneous mutations in outbred populations. With this method we have mapped two spontaneous developmental mutants. In Ciona intestinalis we mapped a short-tail mutation with strong phenotypic similarity to a previously identified mutant in the related species Ciona savignyi. Our bioinformatic approach mapped the mutation to a narrow interval containing a single mutated gene, α-laminin3,4,5, which is the gene previously implicated in C. savignyi. In addition, we mapped a novel genetic mutation disrupting neural tube closure in C. savignyi to a T-type Ca²⁺ channel gene. The high efficiency and unprecedented mapping resolution of our study is a powerful advantage for developmental genetics in Ciona, and may find application in other outbred species.     

中国美利奴羊H-FABP基因多态性及其与部分肉质性状的相关性

利用聚合酶链式反应-单链构象多态（PCR-SSCP）技术检测中国美利奴羊（Ovis aries var. Merino）心型脂肪酸结合蛋白基因（H-FABP）外显子2的单核苷酸多态性（SNPs）和遗传多态性,分析其与肌内脂肪（IMF）含量、肌纤维直径和肌纤维密度的相互关系,为该品种绵羊的分子标记辅助选择提供理论依据。结果显示,H-FABP基因外显子2有AA、AB和BB 3种基因型,AA型和BB型在778位均发生了C缺失,939位均发生了A→G转换,BB型还在789位发生了T→C转换,该突变导致所编码氨基酸发生了缬氨酸→丙氨酸的替换;BB型为IMF的优势基因型,与AB型相比差异显著（P<0.05）,与AA型相比差异极显著（P<0.01）;BB型对肌纤维直径存在负相关。结果提示,中国美利奴羊H-FABP基因外显子2具有多态性,该基因可能是中国美利奴羊肉质性状的主效基因,或者与控制肉质性状的主效基因相连锁。     

Pathogenicity and Interaction of Three Nematode Species on Six Bermudagrasses

The ring nematode (Criconemoides ornatus), stunt nematode (Tylenchorhynchus martini), and sting nematode (Belonolaimus Iongicaudatus) reproduced readily on six bermudagrasses (Common, ''U-3'', ''Tufcote'', ''Continental'', ''Tiffine'', and ''Tifdwarf''). Populations of a single nematode species influenced the population development of a second and third parasitic nematode species on a particular host plant. Activity of most nematodes adversely affected reproduction of other nematode species in mixed cultures. Generally, the number of fibrous roots produced by plants decreased as the number of nematode species in the treatments increased. Tifdwarf bermudagrass appeared to be more tolerant to C. ornatus and T. martini than other grasses tested.     

Editor's choice: Dissection of the Octoploid Strawberry Genome by Deep Sequencing of the Genomes of Fragaria Species

Cultivated strawberry (Fragaria x ananassa) is octoploid and shows allogamous behaviour. The present study aims at dissecting this octoploid genome through comparison with its wild relatives, F. iinumae, F. nipponica, F. nubicola, and F. orientalis by de novo whole-genome sequencing on an Illumina and Roche 454 platforms. The total length of the assembled Illumina genome sequences obtained was 698 Mb for F. x ananassa, and ∼200 Mb each for the four wild species. Subsequently, a virtual reference genome termed FANhybrid_r1.2 was constructed by integrating the sequences of the four homoeologous subgenomes of F. x ananassa, from which heterozygous regions in the Roche 454 and Illumina genome sequences were eliminated. The total length of FANhybrid_r1.2 thus created was 173.2 Mb with the N50 length of 5137 bp. The Illumina-assembled genome sequences of F. x ananassa and the four wild species were then mapped onto the reference genome, along with the previously published F. vesca genome sequence to establish the subgenomic structure of F. x ananassa. The strategy adopted in this study has turned out to be successful in dissecting the genome of octoploid F. x ananassa and appears promising when applied to the analysis of other polyploid plant species.     

Natural variation in the genes responsible for maturity loci E1, E2, E3 and E4 in soybean

Background and Aims

The timing of flowering has a direct impact on successful seed production in plants. Flowering of soybean (Glycine max) is controlled by several E loci, and previous studies identified the genes responsible for the flowering loci E1, E2, E3 and E4. However, natural variation in these genes has not been fully elucidated. The aims of this study were the identification of new alleles, establishment of allele diagnoses, examination of allelic combinations for adaptability, and analysis of the integrated effect of these loci on flowering.

Methods

The sequences of these genes and their flanking regions were determined for 39 accessions by primer walking. Systematic discrimination among alleles was performed using DNA markers. Genotypes at the E1–E4 loci were determined for 63 accessions covering several ecological types using DNA markers and sequencing, and flowering times of these accessions at three sowing times were recorded.

Key Results

A new allele with an insertion of a long interspersed nuclear element (LINE) at the promoter of the E1 locus (e1-re) was identified. Insertion and deletion of 36 bases in the eighth intron (E2-in and E2-dl) were observed at the E2 locus. Systematic discrimination among the alleles at the E1–E3 loci was achieved using PCR-based markers. Allelic combinations at the E1–E4 loci were found to be associated with ecological types, and about 62–66 % of variation of flowering time could be attributed to these loci.

Conclusions

The study advances understanding of the combined roles of the E1–E4 loci in flowering and geographic adaptation, and suggests the existence of unidentified genes for flowering in soybean,     

Identification of Recombination and Positively Selected Genes in Brucella

Brucella is a facultative intracellular bacterium belongs to the class alpha proteobacteria. It causes zoonotic disease brucellosis to wide range of animals. Brucella species are highly conserved in nucleotide level.Here, we employed a comparative genomics approach to examine the role of homologous recombination and positive selection in the evolution of Brucella. For the analysis, we have selected 19 complete genomes from 8 species of Brucella. Among the 1599 core genome predicted, 24 genes were showing signals of recombination but no significant breakpoint was found. The analysis revealed that recombination events are less frequent and the impact of recombination occurred is negligible on the evolution of Brucella. This leads to the view that Brucella is clonally evolved. On other hand, 56 genes (3.5 % of core genome) were showing signals of positive selection. Results suggest that natural selection plays an important role in the evolution of Brucella. Some of the genes that are responsible for the pathogenesis of Brucella were found positively selected, presumably due to their role in avoidance of the host immune system.     

Restriction Fragment Length Polymorphism Separates Species of the Xiphinema americanum Group

The Xiphinema americanum group of species is responsible for vectoring several important virus diseases to perennial crops. Variability of transmission of viruses by different species, and difficulties in separating species by morphometric measurements alone, make it essential to reassess the taxonomic position of several species in the group. The measurement of DNA sequence variability is a sensitive assay that can re-evaluate the separation of species and populations from each other. This study describes how an RFLP approach, in which the restriction sites in transcribed spacers of ribosomal repeats were detected, confirmed the separation of 16 populations of these species into X. americanum, X. rivesi, X. pacificum, and X. bricolensis.     

Three Echinostome Species from Wild Birds in the Republic of Korea

Three echinostome species, i.e., Patagifer bilobus, Petasiger neocomense, and Saakotrema metatestis, are newly recorded in the trematode fauna of the Republic of Korea. They were recovered from 3 species of migratory birds (Platalea minor, Podiceps cristatus, and Egretta garzetta), which were donated by the Wildlife Center of Chungbuk (WCC) and the Conservation Genome Resource Bank for Korean Wildlife (CGRB). Only 1 P. bilobus specimen was recovered from the intestine of a black-faced spoonbill (P. minor), and characterized by the bilobed head crown with a deep dorsal incision and 54 collar spines. Twenty P. neocomense were recovered from the intestine of a great crested grebe (P. cristatus), and they had a well-developed head crown with 19 spines and 2 testes obliquely located at the posterior middle of the body. Total 70 S. metatestis were collected from the bursa of Fabricius of 1 little egret (E. garzetta). It is characterized by stout tegumental spines covered in the entire leaf-shaped body, posterior extension of the uterus, presence of the uroproct and a well-developed head crown with 12 pairs of collar spines on each side. By the present study, these 3 echinostome species are newly added to the trematode fauna in Korea.     

Fine-Scale Heterogeneity in Crossover Rate in the garnet-scalloped Region of the Drosophila melanogaster X Chromosome

Homologous recombination affects myriad aspects of genome evolution, from standing levels of nucleotide diversity to the efficacy of natural selection. Rates of crossing over show marked variability at all scales surveyed, including species-, population-, and individual-level differences. Even within genomes, crossovers are nonrandomly distributed in a wide diversity of taxa. Although intra- and intergenomic heterogeneities in crossover distribution have been documented in Drosophila, the scale and degree of crossover rate heterogeneity remain unclear. In addition, the genetic features mediating this heterogeneity are unknown. Here we quantify fine-scale heterogeneity in crossover distribution in a 2.1-Mb region of the Drosophila melanogaster X chromosome by localizing crossover breakpoints in 2500 individuals, each containing a single crossover in this specific X chromosome region. We show 90-fold variation in rates of crossing over at a 5-kb scale, place this variation in the context of several aspects of genome evolution, and identify several genetic features associated with crossover rates. Our results shed new light on the scale and magnitude of crossover rate heterogeneity in D. melanogaster and highlight potential features mediating this heterogeneity.