Dataset Information

Transcriptome Analysis in Domesticated Species: Challenges and Strategies.

ABSTRACT: Domesticated species occupy a special place in the human world due to their economic and cultural value. In the era of genomic research, domesticated species provide unique advantages for investigation of diseases and complex phenotypes. RNA sequencing, or RNA-seq, has recently emerged as a new approach for studying transcriptional activity of the whole genome, changing the focus from individual genes to gene networks. RNA-seq analysis in domesticated species may complement genome-wide association studies of complex traits with economic importance or direct relevance to biomedical research. However, RNA-seq studies are more challenging in domesticated species than in model organisms. These challenges are at least in part associated with the lack of quality genome assemblies for some domesticated species and the absence of genome assemblies for others. In this review, we discuss strategies for analyzing RNA-seq data, focusing particularly on questions and examples relevant to domesticated species.

SUBMITTER: Hekman JP

PROVIDER: S-EPMC4756862 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Transcriptome Analysis in Domesticated Species: Challenges and Strategies.

Hekman Jessica P JP Johnson Jennifer L JL Kukekova Anna V AV

Bioinformatics and biology insights 20150101 Suppl 4

Domesticated species occupy a special place in the human world due to their economic and cultural value. In the era of genomic research, domesticated species provide unique advantages for investigation of diseases and complex phenotypes. RNA sequencing, or RNA-seq, has recently emerged as a new approach for studying transcriptional activity of the whole genome, changing the focus from individual genes to gene networks. RNA-seq analysis in domesticated species may complement genome-wide associati ...[more]

PMID: 26917953

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Project description:BACKGROUND:Comparative genomics studies are central in identifying the coding and non-coding elements associated with complex traits, and the functional annotation of genomes is a critical step to decipher the genotype-to-phenotype relationships in livestock animals. As part of the Functional Annotation of Animal Genomes (FAANG) action, the FR-AgENCODE project aimed to create reference functional maps of domesticated animals by profiling the landscape of transcription (RNA-seq), chromatin accessibility (ATAC-seq) and conformation (Hi-C) in species representing ruminants (cattle, goat), monogastrics (pig) and birds (chicken), using three target samples related to metabolism (liver) and immunity (CD4+ and CD8+ T cells). RESULTS:RNA-seq assays considerably extended the available catalog of annotated transcripts and identified differentially expressed genes with unknown function, including new syntenic lncRNAs. ATAC-seq highlighted an enrichment for transcription factor binding sites in differentially accessible regions of the chromatin. Comparative analyses revealed a core set of conserved regulatory regions across species. Topologically associating domains (TADs) and epigenetic A/B compartments annotated from Hi-C data were consistent with RNA-seq and ATAC-seq data. Multi-species comparisons showed that conserved TAD boundaries had stronger insulation properties than species-specific ones and that the genomic distribution of orthologous genes in A/B compartments was significantly conserved across species. CONCLUSIONS:We report the first multi-species and multi-assay genome annotation results obtained by a FAANG project. Beyond the generation of reference annotations and the confirmation of previous findings on model animals, the integrative analysis of data from multiple assays and species sheds a new light on the multi-scale selective pressure shaping genome organization from birds to mammals. Overall, these results emphasize the value of FAANG for research on domesticated animals and reinforces the importance of future meta-analyses of the reference datasets being generated by this community on different species.

Project description:Analyses of high-throughput transcriptome sequences of non-model organisms are based on two main approaches: de novo assembly and genome-guided assembly using mapping to assign reads prior to assembly. Given the limits of mapping reads to a reference when it is highly divergent, as is frequently the case for non-model species, we evaluate whether using blastn would outperform mapping methods for read assignment in such situations (>15% divergence). We demonstrate its high performance by using simulated reads of lengths corresponding to those generated by the most common sequencing platforms, and over a realistic range of genetic divergence (0% to 30% divergence). Here we focus on gene identification and not on resolving the whole set of transcripts (i.e. the complete transcriptome). For simulated datasets, the transcriptome-guided assembly based on blastn recovers 94.8% of genes irrespective of read length at 0% divergence; however, assignment rate of reads is negatively correlated with both increasing divergence level and reducing read lengths. Nevertheless, we still observe 92.6% of recovered genes at 30% divergence irrespective of read length. This analysis also produces a categorization of genes relative to their assignment, and suggests guidelines for data processing prior to analyses of comparative transcriptomics and gene expression to minimize potential inferential bias associated with incorrect transcript assignment. We also compare the performances of de novo assembly alone vs in combination with a transcriptome-guided assembly based on blastn both via simulation and empirically, using data from a cyprinid fish species and from an oak species. For any simulated scenario, the transcriptome-guided assembly using blastn outperforms the de novo approach alone, including when the divergence level is beyond the reach of traditional mapping methods. Combining de novo assembly and a related reference transcriptome for read assignment also addresses the bias/error in contigs caused by the dependence on a related reference alone. Empirical data corroborate these findings when assembling transcriptomes from the two non-model organisms: Parachondrostoma toxostoma (fish) and Quercus pubescens (plant). For the fish species, out of the 31,944 genes known from D. rerio, the guided and de novo assemblies recover respectively 20,605 and 20,032 genes but the performance of the guided assembly approach is much higher for both the contiguity and completeness metrics. For the oak, out of the 29,971 genes known from Vitis vinifera, the transcriptome-guided and de novo assemblies display similar performance, but the new guided approach detects 16,326 genes where the de novo assembly only detects 9,385 genes.

Dataset Information

Transcriptome Analysis in Domesticated Species: Challenges and Strategies.

Publications

Transcriptome Analysis in Domesticated Species: Challenges and Strategies.

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