Genome-wide Sequence Information Research Articles

Enigmatic Nodal and Lefty gene repertoire discrepancy: Latent evolutionary history revealed by vertebrate-wide phylogeny.

Homology in vertebrate body plans is traditionally ascribed to the high-level conservation of regulatory components within the genetic programs governing them, particularly during the "phylotypic stage." However, advancements in embryology and molecular phylogeny have unveiled the dynamic nature of gene repertoires responsible for early development. Notably, the Nodal and Lefty genes, members of the transforming growth factor-beta superfamily producing intercellular signaling molecules and crucial for left-right (L-R) symmetry breaking, exhibit distinctive features within their gene repertoires. These features encompass among-species gene repertoire variations resulting from gene gain and loss, as well as gene conversion. Despite their significance, these features have been largely unexplored in a phylogenetic context, but accumulating genome-wide sequence information is allowing the scrutiny of these features. It has exposed hidden paralogy between Nodal1 and Nodal2 genes resulting from differential gene loss in amniotes. In parallel, the tandem cluster of Lefty1 and Lefty2 genes, which was thought to be confined to mammals, is observed in sharks and rays, with an unexpected phylogenetic pattern. This article provides a comprehensive review of the current understanding of the origins of these vertebrate gene repertoires and proposes a revised nomenclature based on the elucidated history of vertebrate genome evolution.

Inference of a genome-wide protein-coding gene set of the inshore hagfish Eptatretus burgeri

The hagfishes (Myxiniformes) arose from agnathan (jawless vertebrate) lineages and they are one of only two extant cyclostome taxa, together with lampreys (Petromyzontiformes). Even though whole genome sequencing has been achieved for diverse vertebrate taxa, genome-wide sequence information has been highly limited for cyclostomes. Here we sequenced the genome of the inshore hagfish Eptatretus burgeri using DNA extracted from the testis, with a short-read sequencing platform, aiming to reconstruct a high-coverage protein-coding gene catalogue. The obtained genome assembly, scaffolded with mate-pair reads and paired RNA-seq reads, exhibited an N50 scaffold length of 293 Kbp, which allowed the genome-wide prediction of coding genes. This computation resulted in the gene models whose completeness was estimated at the complete coverage of more than 83 % and the partial coverage of more than 93 % by referring to evolutionarily conserved single-copy orthologs. The high contiguity of the assembly and completeness of the gene models promise a high utility in various comparative analyses including phylogenomics and phylome exploration.

An open science study of ageing in companion dogs.

The Dog Aging Project is a long-term longitudinal study of ageing in tens of thousands of companion dogs. The domestic dog is among the most variable mammal species in terms of morphology, behaviour, risk of age-related disease and life expectancy. Given that dogs share the human environment and have a sophisticated healthcare system but are much shorter-lived than people, they offer a unique opportunity to identify the genetic, environmental and lifestyle factors associated with healthy lifespan. To take advantage of this opportunity, the Dog Aging Project will collect extensive survey data, environmental information, electronic veterinary medical records, genome-wide sequence information, clinicopathology and molecular phenotypes derived from blood cells, plasma and faecal samples. Here, we describe the specific goals and design of the Dog Aging Project and discuss the potential for this open-data, community science study to greatly enhance understanding of ageing in a genetically variable, socially relevant species living in a complex environment.

Genome-wide sequence information reveals recurrent hybridization among diploid wheat wild relatives.

Many conflicting hypotheses regarding the relationships among crops and wild species closely related to wheat (the genera Aegilops, Amblyopyrum, and Triticum) have been postulated. The contribution of hybridization to the evolution of these taxa is intensely discussed. To determine possible causes for this, and provide a phylogeny of the diploid taxa based on genome-wide sequence information, independent data were obtained from genotyping-by-sequencing and a target-enrichment experiment that returned 244 low-copy nuclear loci. The data were analyzed using Bayesian, likelihood and coalescent-based methods. D statistics were used to test if incomplete lineage sorting alone or together with hybridization is the source for incongruent gene trees. Here we present the phylogeny of all diploid species of the wheat wild relatives. We hypothesize that most of the wheat-group species were shaped by a primordial homoploid hybrid speciation event involving the ancestral Triticum and Am. muticum lineages to form all other species except Ae. speltoides. This hybridization event was followed by multiple introgressions affecting all taxa except Triticum. Mostly progenitors of the extant species were involved in these processes, while recent interspecific gene flow seems insignificant. The composite nature of many genomes of wheat-group taxa results in complicated patterns of diploid contributions when these lineages are involved in polyploid formation, which is, for example, the case for tetraploid and hexaploid wheats. Our analysis provides phylogenetic relationships and a testable hypothesis for the genome compositions in the basic evolutionary units within the wheat group of Triticeae.

Transcriptome-Based Prediction of Complex Traits in Maize.

The ability to predict traits from genome-wide sequence information (i.e., genomic prediction) has improved our understanding of the genetic basis of complex traits and transformed breeding practices. Transcriptome data may also be useful for genomic prediction. However, it remains unclear how well transcript levels can predict traits, particularly when traits are scored at different development stages. Using maize (Zea mays) genetic markers and transcript levels from seedlings to predict mature plant traits, we found that transcript and genetic marker models have similar performance. When the transcripts and genetic markers with the greatest weights (i.e., the most important) in those models were used in one joint model, performance increased. Furthermore, genetic markers important for predictions were not close to or identified as regulatory variants for important transcripts. These findings demonstrate that transcript levels are useful for predicting traits and that their predictive power is not simply due to genetic variation in the transcribed genomic regions. Finally, genetic marker models identified only 1 of 14 benchmark flowering-time genes, while transcript models identified 5. These data highlight that, in addition to being useful for genomic prediction, transcriptome data can provide a link between traits and variation that cannot be readily captured at the sequence level.

Genome-wide analysis of replication timing by next-generation sequencing with E/L Repli-seq.

This protocol is an extension to: Nat. Protoc. 6, 870-895 (2014); doi:10.1038/nprot.2011.328; published online 02 June 2011Cycling cells duplicate their DNA content during S phase, following a defined program called replication timing (RT). Early- and late-replicating regions differ in terms of mutation rates, transcriptional activity, chromatin marks and subnuclear position. Moreover, RT is regulated during development and is altered in diseases. Here, we describe E/L Repli-seq, an extension of our Repli-chip protocol. E/L Repli-seq is a rapid, robust and relatively inexpensive protocol for analyzing RT by next-generation sequencing (NGS), allowing genome-wide assessment of how cellular processes are linked to RT. Briefly, cells are pulse-labeled with BrdU, and early and late S-phase fractions are sorted by flow cytometry. Labeled nascent DNA is immunoprecipitated from both fractions and sequenced. Data processing leads to a single bedGraph file containing the ratio of nascent DNA from early versus late S-phase fractions. The results are comparable to those of Repli-chip, with the additional benefits of genome-wide sequence information and an increased dynamic range. We also provide computational pipelines for downstream analyses, for parsing phased genomes using single-nucleotide polymorphisms (SNPs) to analyze RT allelic asynchrony, and for direct comparison to Repli-chip data. This protocol can be performed in up to 3 d before sequencing, and requires basic cellular and molecular biology skills, as well as a basic understanding of Unix and R.

DISMS2: A flexible algorithm for direct proteome- wide distance calculation of LC-MS/MS runs

BackgroundThe classification of samples on a molecular level has manifold applications, from patient classification regarding cancer treatment to phylogenetics for identifying evolutionary relationships between species. Modern methods employ the alignment of DNA or amino acid sequences, mostly not genome-wide but only on selected parts of the genome. Recently proteomics-based approaches have become popular. An established method for the identification of peptides and proteins is liquid chromatography-tandem mass spectrometry (LC-MS/MS). First, protein sequences from MS/MS spectra are identified by means of database searches, given samples with known genome-wide sequence information, then sequence based methods are applied. Alternatively, de novo peptide sequencing algorithms annotate MS/MS spectra and deduce peptide/protein information without a database. A newer approach independent of additional information is to directly compare unidentified tandem mass spectra. The challenge then is to compute the distance between pairwise MS/MS runs consisting of thousands of spectra.MethodsWe present DISMS2, a new algorithm to calculate proteome-wide distances directly from MS/MS data, extending the algorithm compareMS2, an approach that also uses a spectral comparison pipeline.ResultsOur new more flexible algorithm, DISMS2, allows for the choice of the spectrum distance measure and includes different spectra preprocessing and filtering steps that can be tailored to specific situations by parameter optimization.ConclusionsDISMS2 performs well for samples from species with and without database annotation and thus has clear advantages over methods that are purely based on database search.

Whole-genome sequence-based genomic prediction in laying chickens with different genomic relationship matrices to account for genetic architecture

BackgroundWith the availability of next-generation sequencing technologies, genomic prediction based on whole-genome sequencing (WGS) data is now feasible in animal breeding schemes and was expected to lead to higher predictive ability, since such data may contain all genomic variants including causal mutations. Our objective was to compare prediction ability with high-density (HD) array data and WGS data in a commercial brown layer line with genomic best linear unbiased prediction (GBLUP) models using various approaches to weight single nucleotide polymorphisms (SNPs).MethodsA total of 892 chickens from a commercial brown layer line were genotyped with 336 K segregating SNPs (array data) that included 157 K genic SNPs (i.e. SNPs in or around a gene). For these individuals, genome-wide sequence information was imputed based on data from re-sequencing runs of 25 individuals, leading to 5.2 million (M) imputed SNPs (WGS data), including 2.6 M genic SNPs. De-regressed proofs (DRP) for eggshell strength, feed intake and laying rate were used as quasi-phenotypic data in genomic prediction analyses. Four weighting factors for building a trait-specific genomic relationship matrix were investigated: identical weights, −(log10P) from genome-wide association study results, squares of SNP effects from random regression BLUP, and variable selection based weights (known as BLUP|GA). Predictive ability was measured as the correlation between DRP and direct genomic breeding values in five replications of a fivefold cross-validation.ResultsAveraged over the three traits, the highest predictive ability (0.366 ± 0.075) was obtained when only genic SNPs from WGS data were used. Predictive abilities with genic SNPs and all SNPs from HD array data were 0.361 ± 0.072 and 0.353 ± 0.074, respectively. Prediction with −(log10P) or squares of SNP effects as weighting factors for building a genomic relationship matrix or BLUP|GA did not increase accuracy, compared to that with identical weights, regardless of the SNP set used.ConclusionsOur results show that little or no benefit was gained when using all imputed WGS data to perform genomic prediction compared to using HD array data regardless of the weighting factors tested. However, using only genic SNPs from WGS data had a positive effect on prediction ability.

Ignoring Heterozygous Sites Biases Phylogenomic Estimates of Divergence Times: Implications for the Evolutionary History of Microtus Voles

Phylogenetic reconstruction of the evolutionary history of closely related organisms may be difficult because of the presence of unsorted lineages and of a relatively high proportion of heterozygous sites that are usually not handled well by phylogenetic programs. Genomic data may provide enough fixed polymorphisms to resolve phylogenetic trees, but the diploid nature of sequence data remains analytically challenging. Here, we performed a phylogenomic reconstruction of the evolutionary history of the common vole (Microtus arvalis) with a focus on the influence of heterozygosity on the estimation of intraspecific divergence times. We used genome-wide sequence information from 15 voles distributed across the European range. We provide a novel approach to integrate heterozygous information in existing phylogenetic programs by repeated random haplotype sampling from sequences with multiple unphased heterozygous sites. We evaluated the impact of the use of full, partial, or no heterozygous information for tree reconstructions on divergence time estimates. All results consistently showed four deep and strongly supported evolutionary lineages in the vole data. These lineages undergoing divergence processes split only at the end or after the last glacial maximum based on calibration with radiocarbon-dated paleontological material. However, the incorporation of information from heterozygous sites had a significant impact on absolute and relative branch length estimations. Ignoring heterozygous information led to an overestimation of divergence times between the evolutionary lineages of M. arvalis. We conclude that the exclusion of heterozygous sites from evolutionary analyses may cause biased and misleading divergence time estimates in closely related taxa.

Genome-wide BAC-end sequencing of Cucumis melo using two BAC libraries

BackgroundAlthough melon (Cucumis melo L.) is an economically important fruit crop, no genome-wide sequence information is openly available at the current time. We therefore sequenced BAC-ends representing a total of 33,024 clones, half of them from a previously described melon BAC library generated with restriction endonucleases and the remainder from a new random-shear BAC library.ResultsWe generated a total of 47,140 high-quality BAC-end sequences (BES), 91.7% of which were paired-BES. Both libraries were assembled independently and then cross-assembled to obtain a final set of 33,372 non-redundant, high-quality sequences. These were grouped into 6,411 contigs (4.5 Mb) and 26,961 non-assembled BES (14.4 Mb), representing ~4.2% of the melon genome. The sequences were used to screen genomic databases, identifying 7,198 simple sequence repeats (corresponding to one microsatellite every 2.6 kb) and 2,484 additional repeats of which 95.9% represented transposable elements. The sequences were also used to screen expressed sequence tag (EST) databases, revealing 11,372 BES that were homologous to ESTs. This suggests that ~30% of the melon genome consists of coding DNA. We observed regions of microsynteny between melon paired-BES and six other dicotyledonous plant genomes.ConclusionThe analysis of nearly 50,000 BES from two complementary genomic libraries covered ~4.2% of the melon genome, providing insight into properties such as microsatellite and transposable element distribution, and the percentage of coding DNA. The observed synteny between melon paired-BES and six other plant genomes showed that useful comparative genomic data can be derived through large scale BAC-end sequencing by anchoring a small proportion of the melon genome to other sequenced genomes.