Nathaniel K. Jue

Two different high throughput sequencing approaches identify thousands of de novo genomic markers for the genetically depleted Bornean elephant.

High throughput sequencing technologies are being applied to an increasing number of model species with a high-quality reference genome. The application and analyses of whole-genome sequence data in non-model species with no prior genomic information are currently under way. Recent sequencing technologies provide new opportunities for gathering genomic data in natural populations, laying the empirical foundation for future research in the field of conservation and population genomics. Here we present the case study of the Bornean elephant, which is the most endangered subspecies of Asian elephant and exhibits very low genetic diversity. We used two different sequencing platforms, the Roche 454 FLX (shotgun) and Illumina, GAIIx (Restriction site associated DNA, RAD) to evaluate the feasibility of the two methodologies for the discovery of de novo markers (single nucleotide polymorphism, SNPs and microsatellites) using low coverage data. Approximately, 6,683 (shotgun) and 14,724 (RAD) SNPs were detected within our elephant sequence dataset. Genotyping of a representative sample of 194 SNPs resulted in a SNP validation rate of ∼ 83 to 94% and 17% of the loci were polymorphic with a low diversity (H o = 0.057). Different numbers of microsatellites were identified through shotgun (27,226) and RAD (868) techniques. Out of all di-, tri-, and tetra-microsatellite loci, 1,706 loci had sufficient flanking regions (shotgun) while only 7 were found with RAD. All microsatellites were monomorphic in the Bornean but polymorphic in another elephant subspecies. Despite using different sample sizes, and the well known differences in the two platforms used regarding sequence length and throughput, the two approaches showed high validation rate. The approaches used here for marker development in a threatened species demonstrate the utility of high throughput sequencing technologies as a starting point for the development of genomic tools in a non-model species and in particular for a species with low genetic diversity.

Determination of dosage compensation of the mammalian X chromosome by RNA-seq is dependent on analytical approach

BackgroundAn enduring question surrounding sex chromosome evolution is whether effective hemizygosity in the heterogametic sex leads inevitably to dosage compensation of sex-linked genes, and whether this compensation has been observed in a variety of organisms. Incongruence in the conclusions reached in some recent reports has been attributed to different high-throughput approaches to transcriptome analysis. However, recent reports each utilizing RNA-seq to gauge X-linked gene expression relative to autosomal gene expression also arrived at diametrically opposed conclusions regarding X chromosome dosage compensation in mammals.ResultsHere we analyze RNA-seq data from X-monosomic female human and mouse tissues, which are uncomplicated by genes that escape X-inactivation, as well as published RNA-seq data to describe relative X expression (RXE). We find that the determination of RXE is highly dependent upon a variety of computational, statistical and biological assumptions underlying RNA-seq analysis. Parameters implemented in short-read mapping programs, choice of reference genome annotation, expression data distribution, tissue source for RNA and RNA-seq library construction method have profound effects on comparing expression levels across chromosomes.ConclusionsOur analysis shows that the high number of paralogous gene families on the mammalian X chromosome relative to autosomes contributes to the ambiguity in RXE calculations, RNA-seq analysis that takes into account that single- and multi-copy genes are compensated differently supports the conclusion that, in many somatic tissues, the mammalian X is up-regulated compared to the autosomes.

Recent Amplification of the Kangaroo Endogenous Retrovirus, KERV, Limited to the Centromere

ABSTRACT Mammalian retrotransposons, transposable elements that are processed through an RNA intermediate, are categorized as short interspersed elements (SINEs), long interspersed elements (LINEs), and long terminal repeat (LTR) retroelements, which include endogenous retroviruses. The ability of transposable elements to autonomously amplify led to their initial characterization as selfish or junk DNA; however, it is now known that they may acquire specific cellular functions in a genome and are implicated in host defense mechanisms as well as in genome evolution. Interactions between classes of transposable elements may exert a markedly different and potentially more significant effect on a genome than interactions between members of a single class of transposable elements. We examined the genomic structure and evolution of the kangaroo endogenous retrovirus (KERV) in the marsupial genus Macropus. The complete proviral structure of the kangaroo endogenous retrovirus, phylogenetic relationship among relative retroviruses, and expression of this virus in both Macropus rufogriseus and M. eugenii are presented for the first time. In addition, we show the relative copy number and distribution of the kangaroo endogenous retrovirus in the Macropus genus. Our data indicate that amplification of the kangaroo endogenous retrovirus occurred in a lineage-specific fashion, is restricted to the centromeres, and is not correlated with LINE depletion. Finally, analysis of KERV long terminal repeat sequences using massively parallel sequencing indicates that the recent amplification in M. rufogriseus is likely due to duplications and concerted evolution rather than a high number of independent insertion events.

Rapid Evolutionary Rates and Unique Genomic Signatures Discovered in the First Reference Genome for the Southern Ocean Salp, Salpa thompsoni (Urochordata, Thaliacea).

A preliminary genome sequence has been assembled for the Southern Ocean salp, Salpa thompsoni (Urochordata, Thaliacea). Despite the ecological importance of this species in Antarctic pelagic food webs and its potential role as an indicator of changing Southern Ocean ecosystems in response to climate change, no genomic resources are available for S. thompsoni or any closely related urochordate species. Using a multiple-platform, multiple-individual approach, we have produced a 318,767,936-bp genome sequence, covering >50% of the estimated 602 Mb (±173 Mb) genome size for S. thompsoni. Using a nonredundant set of predicted proteins, >50% (16,823) of sequences showed significant homology to known proteins and ∼38% (12,151) of the total protein predictions were associated with Gene Ontology functional information. We have generated 109,958 SNP variant and 9,782 indel predictions for this species, serving as a resource for future phylogenomic and population genetic studies. Comparing the salp genome to available assemblies for four other urochordates, Botryllus schlosseri, Ciona intestinalis, Ciona savignyi and Oikopleura dioica, we found that S. thompsoni shares the previously estimated rapid rates of evolution for these species. High mutation rates are thus independent of genome size, suggesting that rates of evolution >1.5 times that observed for vertebrates are a broad taxonomic characteristic of urochordates. Tests for positive selection implemented in PAML revealed a small number of genes with sites undergoing rapid evolution, including genes involved in ribosome biogenesis and metabolic and immune process that may be reflective of both adaptation to polar, planktonic environments as well as the complex life history of the salps. Finally, we performed an initial survey of small RNAs, revealing the presence of known, conserved miRNAs, as well as novel miRNA genes; unique piRNAs; and mature miRNA signatures for varying developmental stages. Collectively, these resources provide a genomic foundation supporting S. thompsoni as a model species for further examination of the exceptional rates and patterns of genomic evolution shown by urochordates. Additionally, genomic data will allow for the development of molecular indicators of key life history events and processes and afford new understandings and predictions of impacts of climate change on this key species of Antarctic pelagic ecosystems.

From Shelf to Shelf: Assessing Historical and Contemporary Genetic Differentiation and Connectivity across the Gulf of Mexico in Gag, Mycteroperca microlepis

Describing patterns of connectivity among populations of species with widespread distributions is particularly important in understanding the ecology and evolution of marine species. In this study, we examined patterns of population differentiation, migration, and historical population dynamics using microsatellite and mitochondrial loci to test whether populations of the epinephelid fish, Gag, Mycteroperca microlepis, an important fishery species, are genetically connected across the Gulf of Mexico and if so, whether that connectivity is attributable to either contemporary or historical processes. Populations of Gag on the Campeche Bank and the West Florida Shelf show significant, but low magnitude, differentiation. Time since divergence/expansion estimates associated with historical population dynamics indicate that any population or spatial expansions indicated by population genetics would have likely occurred in the late Pleistocene. Using coalescent-based approaches, we find that the best model for explaining observed spatial patterns of contemporary genetic variation is one of asymmetric gene flow, with movement from Campeche Bank to the West Florida Shelf. Both estimated migration rates and ecological data support the hypothesis that Gag populations throughout the Gulf of Mexico are connected via present day larval dispersal. Demonstrating this greatly expanded scale of connectivity for Gag highlights the influence of “ghost” populations (sensu Beerli) on genetic patterns and presents a critical consideration for both fisheries management and conservation of this and other species with similar genetic patterns.

Tissue-Specific Transcriptome for Poeciliopsis prolifica Reveals Evidence for Genetic Adaptation Related to the Evolution of a Placental Fish

The evolution of the placenta is an excellent model to examine the evolutionary processes underlying adaptive complexity due to the recent, independent derivation of placentation in divergent animal lineages. In fishes, the family Poeciliidae offers the opportunity to study placental evolution with respect to variation in degree of post-fertilization maternal provisioning among closely related sister species. In this study, we present a detailed examination of a new reference transcriptome sequence for the live-bearing, matrotrophic fish, Poeciliopsis prolifica, from multiple-tissue RNA-seq data. We describe the genetic components active in liver, brain, late-stage embryo, and the maternal placental/ovarian complex, as well as associated patterns of positive selection in a suite of orthologous genes found in fishes. Results indicate the expression of many signaling transcripts, “non-coding” sequences and repetitive elements in the maternal placental/ovarian complex. Moreover, patterns of positive selection in protein sequence evolution were found associated with live-bearing fishes, generally, and the placental P. prolifica, specifically, that appear independent of the general live-bearer lifestyle. Much of the observed patterns of gene expression and positive selection are congruent with the evolution of placentation in fish functionally converging with mammalian placental evolution and with the patterns of rapid evolution facilitated by the teleost-specific whole genome duplication event.