Estimation of linkage disequilibrium and interspecific gene flow in Ficedula flycatchers by a newly developed 50k single‐nucleotide polymorphism array

With the access to draft genome sequence assemblies and whole‐genome resequencing data from population samples, molecular ecology studies will be able to take truly genome‐wide approaches. This now applies to an avian model system in ecological and evolutionary research: Old World flycatchers of the genus Ficedula, for which we recently obtained a 1.1 Gb collared flycatcher genome assembly and identified 13 million single‐nucleotide polymorphism (SNP)s in population resequencing of this species and its sister species, pied flycatcher. Here, we developed a custom 50K Illumina iSelect flycatcher SNP array with markers covering 30 autosomes and the Z chromosome. Using a number of selection criteria for inclusion in the array, both genotyping success rate and polymorphism information content (mean marker heterozygosity = 0.41) were high. We used the array to assess linkage disequilibrium (LD) and hybridization in flycatchers. Linkage disequilibrium declined quickly to the background level at an average distance of 17 kb, but the extent of LD varied markedly within the genome and was more than 10‐fold higher in ‘genomic islands’ of differentiation than in the rest of the genome. Genetic ancestry analysis identified 33 F₁ hybrids but no later‐generation hybrids from sympatric populations of collared flycatchers and pied flycatchers, contradicting earlier reports of backcrosses identified from much fewer number of markers. With an estimated divergence time as recently as <1 Ma, this suggests strong selection against F₁ hybrids and unusually rapid evolution of reproductive incompatibility in an avian system.     

Genetic mapping in a natural population of collared flycatchers (Ficedula albicollis): conserved synteny but gene order rearrangements on the avian Z chromosome

Data from completely sequenced genomes are likely to open the way for novel studies of the genetics of nonmodel organisms, in particular when it comes to the identification and analysis of genes responsible for traits that are under selection in natural populations. Here we use the draft sequence of the chicken genome as a starting point for linkage mapping in a wild bird species, the collared flycatcher - one of the most well-studied avian species in ecological and evolutionary research. A pedigree of 365 flycatchers was established and genotyped for single nucleotide polymorphisms in 23 genes selected from (and spread over most of) the chicken Z chromosome. All genes were also found to be located on the Z chromosome in the collared flycatcher, confirming conserved synteny at the level of gene content across distantly related avian lineages. This high degree of conservation mimics the situation seen for the mammalian X chromosome and may thus be a general feature in sex chromosome evolution, irrespective of whether there is male or female heterogamety. Alternatively, such unprecedented chromosomal conservation may be characteristic of most chromosomes in avian genome evolution. However, several internal rearrangements were observed, meaning that the transfer of map information from chicken to nonmodel bird species cannot always assume conserved gene orders. Interestingly, the rate of recombination on the Z chromosome of collared flycatchers was only approximately 50% that of chicken, challenging the widely held view that birds generally have high recombination rates.     

A gene-based genetic linkage map of the collared flycatcher (Ficedula albicollis) reveals extensive synteny and gene-order conservation during 100 million years of avian evolution

By taking advantage of a recently developed reference marker set for avian genome analysis we have constructed a gene-based genetic map of the collared flycatcher, an important "ecological model" for studies of life-history evolution, sexual selection, speciation, and quantitative genetics. A pedigree of 322 birds from a natural population was genotyped for 384 single nucleotide polymorphisms (SNPs) from 170 protein-coding genes and 71 microsatellites. Altogether, 147 gene markers and 64 microsatellites form 33 linkage groups with a total genetic distance of 1787 cM. Male recombination rates are, on average, 22% higher than female rates (total distance 1982 vs. 1627 cM). The ability to anchor the collared flycatcher map with the chicken genome via the gene-based SNPs revealed an extraordinary degree of both synteny and gene-order conservation during avian evolution. The great majority of chicken chromosomes correspond to a single linkage group in collared flycatchers, with only a few cases of inter- and intrachromosomal rearrangements. The rate of chromosomal diversification, fissions/fusions, and inversions combined is thus considerably lower in birds (0.05/MY) than in mammals (0.6-2.0/MY). A dearth of repeat elements, known to promote chromosomal breakage, in avian genomes may contribute to their stability. The degree of genome stability is likely to have important consequences for general evolutionary patterns and may explain, for example, the comparatively slow rate by which genetic incompatibility among lineages of birds evolves.     

A HIGH‐DENSITY SCAN OF THE Z CHROMOSOME IN FICEDULA FLYCATCHERS REVEALS CANDIDATE LOCI FOR DIVERSIFYING SELECTION

Theoretical and empirical data suggest that genes located on sex chromosomes may play an important role both for sexually selected traits and for traits involved in the build‐up of hybrid incompatibilities. We investigated patterns of genetic variation in 73 genes located on the Z chromosomes of two species of the flycatcher genus Ficedula, the pied flycatcher and the collared flycatcher. Sequence data were evaluated for signs of selection potentially related to genomic differentiation in these young sister species, which hybridize despite reduced fitness of hybrids. Seven loci were significantly more divergent between the two species than expected under neutrality and they also displayed reduced nucleotide diversity, consistent with having been influenced by directional selection. Two of the detected candidate regions contain genes that are associated with plumage coloration in birds. Plumage characteristics play an important role in species recognition in these flycatchers suggesting that the detected genes may have been involved in the evolution of sexual isolation between the species.     

Genomics of natural bird populations: a gene-based set of reference markers evenly spread across the avian genome

Although there is growing interest to take genomics into the complex realms of natural populations, there is a general shortage of genomic resources and tools available for wild species. This applies not at least to birds, for which genomic approaches should be helpful to questions such as adaptation, speciation and population genetics. In this study, we describe a genome-wide reference set of conserved avian gene markers, broadly applicable across birds. By aligning protein-coding sequences from the recently assembled chicken genome with orthologous sequences in zebra finch, we identified particularly conserved exonic regions flanking introns of suitable size for subsequent amplification and sequencing. Primers were designed for 242 gene markers evenly distributed across the chicken genome, with a mean inter-marker interval of 4.2 Mb. Between 78% and 93% of the markers amplified a specific product in five species tested (chicken, peregrine falcon, collared flycatcher, great reed warbler and blue tit). Two hundred markers were sequenced in collared flycatcher, yielding a total of 122.41 kb of genomic DNA sequence (12096 bp coding sequence and 110 314 bp noncoding). Intron size of collared flycatcher and chicken was highly correlated, as was GC content. A polymorphism screening using these markers in a panel of 10 unrelated collared flycatchers identified 871 single nucleotide polymorphisms (pi = 0.0029) and 33 indels (mainly very short). Avian genome characteristics such as uniform genome size and low rate of syntenic rearrangements suggest that this marker set will find broad utility as a genome-wide reference resource for molecular ecological and population genomic analysis of birds. We envision that it will be particularly useful for obtaining large-scale orthologous targets in different species--important in, for instance, phylogenetics--and for large-scale identification of evenly distributed single nucleotide polymorphisms needed in linkage mapping or in studies of gene flow and hybridization.     

家蚕LTR逆转录转座子的鉴定、分类及系统发育分析

转座子是真核生物基因组的重要组成成分。为了研究家蚕Bombyx mori长末端重复序列 (long terminal repeat, LTR)逆转录转座子的分类及进化, 本研究采用de novo预测和同源性搜索相结合的方法, 在家蚕基因组中共鉴定出了38个LTR逆转录转座子家族, 序列长度占整个基因组的0.64%, 远小于先前预测的11.8%, 其中有6个家族为本研究的新发现。38个家族中, 26个家族有表达序列标签 (expression sequence tag, EST)证据, 表明这些家族具有潜在的活性。对有EST证据的6个家族和没有EST证据的5个家族用RT-PCR进行了组织表达谱实验, 结果表明这11个家族在一些组织中有表达, 这进一步证实了这些家族具有转录活性, 基于此我们推测家蚕中大部分的LTR逆转录转座子家族很可能具有潜在活性。对转座子的插入时间进行估计, 结果表明绝大部分元件都是最近1百万年内插入到家蚕基因组中的。我们还比较了黑腹果蝇Drosophila melanogaster、 冈比亚按蚊Anopheles gambiae和家蚕B. mori中Ty3/Gypsy超家族分支的差异, 结果表明不同枝在不同昆虫中有着不同的扩张。家蚕中LTR逆转录转座子的鉴定和系统分析有助于我们理解逆转录转座子在昆虫进化中的作用。     

The influence of transposable elements on animal colouration

Transposable elements (TEs) are mobile genetic sequences present within host genomes. TEs can contribute to the evolution of host traits, since transposition is mutagenic and TEs often contain host regulatory and protein coding sequences. We review cases where TEs influence animal colouration, reporting major patterns and outstanding questions. TE-induced colouration phenotypes typically arise via introduction of novel regulatory sequences and splice sites, affecting pigment cell development or pigment synthesis. We discuss if particular TE types may be more frequently involved in the evolution of colour variation in animals, given that examples involving long terminal repeat (LTR) elements appear to dominate. Currently, examples of TE-induced colouration phenotypes in animals mainly concern model and domesticated insect and mammal species. However, several influential recent examples, coupled with increases in genome sequencing, suggest cases reported from wild species will increase considerably.     

Analysis of Transposable Elements in the Genome of Asparagus officinalis from High Coverage Sequence Data

Asparagus officinalis is an economically and nutritionally important vegetable crop that is widely cultivated and is used as a model dioecious species to study plant sex determination and sex chromosome evolution. To improve our understanding of its genome composition, especially with respect to transposable elements (TEs), which make up the majority of the genome, we performed Illumina HiSeq2000 sequencing of both male and female asparagus genomes followed by bioinformatics analysis. We generated 17 Gb of sequence (12×coverage) and assembled them into 163,406 scaffolds with a total cumulated length of 400 Mbp, which represent about 30% of asparagus genome. Overall, TEs masked about 53% of the A. officinalis assembly. Majority of the identified TEs belonged to LTR retrotransposons, which constitute about 28% of genomic DNA, with Ty1/copia elements being more diverse and accumulated to higher copy numbers than Ty3/gypsy. Compared with LTR retrotransposons, non-LTR retrotransposons and DNA transposons were relatively rare. In addition, comparison of the abundance of the TE groups between male and female genomes showed that the overall TE composition was highly similar, with only slight differences in the abundance of several TE groups, which is consistent with the relatively recent origin of asparagus sex chromosomes. This study greatly improves our knowledge of the repetitive sequence construction of asparagus, which facilitates the identification of TEs responsible for the early evolution of plant sex chromosomes and is helpful for further studies on this dioecious plant.     

Evolutionary dynamics of retrotransposons following autopolyploidy in the Buckler Mustard species complex

Long terminal repeat retrotransposons (LTR‐RTs) represent a major fraction of plant genomes, but processes leading to transposition bursts remain elusive. Polyploidy expectedly leads to LTR‐RT proliferation, as the merging of divergent diploids provokes a genome shock activating LTR‐RTs and/or genetic redundancy supports the accumulation of active LTR‐RTs through relaxation of selective constraints. Available evidence supports interspecific hybridization as the main trigger of genome dynamics, but few studies have addressed the consequences of intraspecific polyploidy (i.e. autopolyploidy), where the genome shock is expectedly minimized. The dynamics of LTR‐RTs was thus here evaluated through low coverage 454 sequencing of three closely related diploid progenitors and three independent autotetraploids from the young Biscutella laevigata species complex. Genomes from this early diverging Brassicaceae lineage presented a minimum of 40% repeats and a large diversity of transposable elements. Differential abundances and patterns of sequence divergence among genomes for 37 LTR‐RT families revealed contrasted dynamics during species diversification. Quiescent LTR‐RT families with limited genetic variation among genomes were distinguished from active families (37.8%) having proliferated in specific taxa. Specific families proliferated in autopolyploids only, but most transpositionally active families in polyploids were also differentiated among diploids. Low expression levels of transpositionally active LTR‐RT families in autopolyploids further supported that genome shock and redundancy are non‐mutually exclusive triggers of LTR‐RT proliferation. Although reputed stable, autopolyploid genomes show LTR‐RT fractions presenting analogies with polyploids between widely divergent genomes.     

Song similarity predicts hybridization in flycatchers

Given that population divergence in sexual signals is an important prerequisite for reproductive isolation, a key prediction is that cases of signal convergence should lead to hybridization. However, empirical studies that quantitatively demonstrate links between phenotypic characters of individuals and their likelihood to hybridize are rare. Here we show that song convergence between sympatric pied (Ficedula hypoleuca) and collared flycatchers (F. albicollis) influence social and sexual interactions between the two species. In sympatry, the majority of male pied flycatchers (65%) include various parts of collared flycatcher song in their song repertoire (but not vice versa). Playback experiments on male interactions demonstrate that male collared flycatchers respond similarly to this 'mixed' song as to conspecific song. Long-term data on pairing patterns show that males singing a converged song attract females of the other species: female collared flycatchers only pair with male pied flycatchers if the males sing the mixed song type. From the perspective of a male pied flycatcher, singing a mixed song type is associated with 30% likelihood of hybridization. This result, combined with our estimates of the frequency of mixed singers, accurately predicts the observed occurrence of hybridization among male pied flycatchers in our study populations (20.45% of 484 pairs; predicted 19.5%). Our results support the suggestion that song functions as the most important prezygotic isolation mechanism in many birds.     

What makes transposable elements move in the Drosophila genome?

Transposable elements (TEs), by their capacity of moving and inducing mutations in the genome, are considered important drivers of species evolution. The successful invasions of TEs in genomes, despite their mutational properties, are an apparent paradox. TEs' transposition is usually strongly regulated to low value, but in some cases these elements can also show high transposition rates, which has been associated sometimes to changes in environmental conditions. It is evident that factors susceptible to induce transpositions in natural populations contribute to TE perpetuation. Different factors were proposed as causative agents of TE mobilization in a wide range of organisms: biotic and abiotic stresses, inter- and intraspecific crosses and populational factors. However, there is no clear evidence of the factors capable of inducing TE mobilization in Drosophila, and data on laboratory stocks show contradictory results. The aim of this review is to have an update critical revision about mechanisms promoting transposition of TEs in Drosophila, and to provide to the readers a global vision of the dynamics of these genomic elements in the Drosophila genome.     

Updating of transposable element annotations from large wheat genomic sequences reveals diverse activities and gene associations

Triticeae species (including wheat, barley and rye) have huge and complex genomes due to polyploidization and a high content of transposable elements (TEs). TEs are known to play a major role in the structure and evolutionary dynamics of Triticeae genomes. During the last 5 years, substantial stretches of contiguous genomic sequence from various species of Triticeae have been generated, making it necessary to update and standardize TE annotations and nomenclature. In this study we propose standard procedures for these tasks, based on structure, nucleic acid and protein sequence homologies. We report statistical analyses of TE composition and distribution in large blocks of genomic sequences from wheat and barley. Altogether, 3.8 Mb of wheat sequence available in the databases was analyzed or re-analyzed, and compared with 1.3 Mb of re-annotated genomic sequences from barley. The wheat sequences were relatively gene-rich (one gene per 23.9 kb), although wheat gene-derived sequences represented only 7.8% (159 elements) of the total, while the remainder mainly comprised coding sequences found in TEs (54.7%, 751 elements). Class I elements [mainly long terminal repeat (LTR) retrotransposons] accounted for the major proportion of TEs, in terms of sequence length as well as element number (83.6% and 498, respectively). In addition, we show that the gene-rich sequences of wheat genome A seem to have a higher TE content than those of genomes B and D, or of barley gene-rich sequences. Moreover, among the various TE groups, MITEs were most often associated with genes: 43.1% of MITEs fell into this category. Finally, the TRIM and copia elements were shown to be the most active TEs in the wheat genome. The implications of these results for the evolution of diploid and polyploid wheat species are discussed. Electronic Supplementary Material Supplementary material is available for this article at     

Illumina TruSeq Synthetic Long-Reads Empower De Novo Assembly and Resolve Complex,Highly-Repetitive Transposable Elements

High-throughput DNA sequencing technologies have revolutionized genomic analysis, including the de novo assembly of whole genomes. Nevertheless, assembly of complex genomes remains challenging, in part due to the presence of dispersed repeats which introduce ambiguity during genome reconstruction. Transposable elements (TEs) can be particularly problematic, especially for TE families exhibiting high sequence identity, high copy number, or complex genomic arrangements. While TEs strongly affect genome function and evolution, most current de novo assembly approaches cannot resolve long, identical, and abundant families of TEs. Here, we applied a novel Illumina technology called TruSeq synthetic long-reads, which are generated through highly-parallel library preparation and local assembly of short read data and which achieve lengths of 1.5–18.5 Kbp with an extremely low error rate (0.03% per base). To test the utility of this technology, we sequenced and assembled the genome of the model organism Drosophila melanogaster (reference genome strain y; cn, bw, sp) achieving an N50 contig size of 69.7 Kbp and covering 96.9% of the euchromatic chromosome arms of the current reference genome. TruSeq synthetic long-read technology enables placement of individual TE copies in their proper genomic locations as well as accurate reconstruction of TE sequences. We entirely recovered and accurately placed 4,229 (77.8%) of the 5,434 annotated transposable elements with perfect identity to the current reference genome. As TEs are ubiquitous features of genomes of many species, TruSeq synthetic long-reads, and likely other methods that generate long-reads, offer a powerful approach to improve de novo assemblies of whole genomes.     

Novel Transposable Elements in Solanaceae: Evolutionary Relationships among Tnt1-related Sequences in Wild Petunia Species

Transposable elements (TEs) are widespread in eukaryotic genomes. The diversity and abundance of TEs are highly variable among species and may correspond to particular relationships between a species and the elements in its genome. There are often many TE families within a single genome; thus, the amplification of one TE family may influence the amplification of other families. LTR retrotransposons (LTR-RTs) are extremely abundant in flowering plants, and Tnt1 is one of the most well known. First characterized in tobacco, Tnt1-related sequences have since been reported in other genera of Solanaceae. In this study, we investigated the profile of Tnt1-related sequences among the species of three Solanaceae genera through genomic amplification and the cloning of partial sequences. The analysis of these sequences revealed high levels of diversity and showed that the sequences are not as closely related to Tnt1 as had been previously hypothesized. The classification of the sequences yielded ten possible families of LTR-RTs, which are, in addition to Tnt1, all members of the Tork clade within the Copia superfamily. However, the sequences did not follow the phylogeny of the species and were not homogeneously distributed. One family includes only sequences of taxa that inhabit dry areas. These findings were consistent with previous suggestions of an early association of Tnt1-related elements with the evolution of several Solanaceae species.     

The evolution of retrotransposon regulatory regions and its consequences on the Drosophila melanogaster and Homo sapiens host genomes

It has now been established that transposable elements (TEs) make up a variable, but significant proportion of the genomes of all organisms, from Bacteria to Vertebrates. However, in addition to their quantitative importance, there is increasing evidence that TEs also play a functional role within the genome. In particular, TE regulatory regions can be viewed as a large pool of potential promoter sequences for host genes. Studying the evolution of regulatory region of TEs in different genomic contexts is therefore a fundamental aspect of understanding how a genome works. In this paper, we first briefly describe what is currently known about the regulation of TE copy number and activity in genomes, and then focus on TE regulatory regions and their evolution. We restrict ourselves to retrotransposons, which are the most abundant class of eukaryotic TEs, and analyze their evolution and the subsequent consequences for host genomes. Particular attention is paid to much-studied representatives of the Vertebrates and Invertebrates, Homo sapiens and Drosophila melanogaster, respectively, for which high quality sequenced genomes are available.     

A Snapshot of Histone Modifications within Transposable Elements in Drosophila Wild Type Strains

Transposable elements (TEs) are a major source of genetic variability in genomes, creating genetic novelty and driving genome evolution. Analysis of sequenced genomes has revealed considerable diversity in TE families, copy number, and localization between different, closely related species. For instance, although the twin species Drosophila melanogaster and D. simulans share the same TE families, they display different amounts of TEs. Furthermore, previous analyses of wild type derived strains of D. simulans have revealed high polymorphism regarding TE copy number within this species. Several factors may influence the diversity and abundance of TEs in a genome, including molecular mechanisms such as epigenetic factors, which could be a source of variation in TE success. In this paper, we present the first analysis of the epigenetic status of four TE families (roo, tirant, 412 and F) in seven wild type strains of D. melanogaster and D. simulans. Our data shows intra- and inter-specific variations in the histone marks that adorn TE copies. Our results demonstrate that the chromatin state of common TEs varies among TE families, between closely related species and also between wild type strains.     

Tempo and Mode of Transposable Element Activity in Drosophila

The evolutionary dynamics of transposable element (TE) insertions have been of continued interest since TE activity has important implications for genome evolution and adaptation. Here, we infer the transposition dynamics of TEs by comparing their abundance in natural D. melanogaster and D. simulans populations. Sequencing pools of more than 550 South African flies to at least 320-fold coverage, we determined the genome wide TE insertion frequencies in both species. We suggest that the predominance of low frequency insertions in the two species (>80% of the insertions have a frequency <0.2) is probably due to a high activity of more than 58 families in both species. We provide evidence for 50% of the TE families having temporally heterogenous transposition rates with different TE families being affected in the two species. While in D. melanogaster retrotransposons were more active, DNA transposons showed higher activity levels in D. simulans. Moreover, we suggest that LTR insertions are mostly of recent origin in both species, while DNA and non-LTR insertions are older and more frequently vertically transmitted since the split of D. melanogaster and D. simulans. We propose that the high TE activity is of recent origin in both species and a consequence of the demographic history, with habitat expansion triggering a period of rapid evolution.     

Large-scale discovery of insertion hotspots and preferential integration sites of human transposed elements

Throughout evolution, eukaryotic genomes have been invaded by transposable elements (TEs). Little is known about the factors leading to genomic proliferation of TEs, their preferred integration sites and the molecular mechanisms underlying their insertion. We analyzed hundreds of thousands nested TEs in the human genome, i.e. insertions of TEs into existing ones. We first discovered that most TEs insert within specific ‘hotspots’ along the targeted TE. In particular, retrotransposed Alu elements contain a non-canonical single nucleotide hotspot for insertion of other Alu sequences. We next devised a method for identification of integration sequence motifs of inserted TEs that are conserved within the targeted TEs. This method revealed novel sequences motifs characterizing insertions of various important TE families: Alu, hAT, ERV1 and MaLR. Finally, we performed a global assessment to determine the extent to which young TEs tend to nest within older transposed elements and identified a 4-fold higher tendency of TEs to insert into existing TEs than to insert within non-TE intergenic regions. Our analysis demonstrates that TEs are highly biased to insert within certain TEs, in specific orientations and within specific targeted TE positions. TE nesting events also reveal new characteristics of the molecular mechanisms underlying transposition.