DOI: 10.5061/dryad.2v3h4
title: Data from: Optimisation of next generation sequencing transcriptome annotation for species lacking sequenced genomes

description: Next generation sequencing methods, such as RNA-seq, have permitted the
 exploration of gene expression in a range of organisms which have been
 studied in ecological contexts but lack a sequenced genome. However, the
 efficacy and accuracy of RNA-seq annotation methods using reference
 genomes from related species have yet to be robustly characterised. Here
 we conduct a comprehensive power analysis employing RNA-seq data from
 Drosophila melanogaster in conjunction with 11 additional genomes from
 related Drosophila species to compare annotation methods and quantify the
 impact of evolutionary divergence between transcriptome and the reference
 genome. Our analyses demonstrate that, regardless of the level of sequence
 divergence, direct genome mapping, where transcript short reads are
 aligned directly to the reference genome, significantly outperforms the
 widely used de novo and guided assembly-based methods in both the quantity
 and accuracy of gene detection. Our analysis also reveals that direct
 genome mapping recovers a more representative profile of Gene Ontology
 functional categories, which are often used to interpret emergent patterns
 in genome-wide expression analyses. Lastly, analysis of available primate
 RNA-seq data demonstrates the applicability of our observations across
 diverse taxa. Our quantification of annotation accuracy and reduced gene
 detection associated with sequence divergence thus provide empirically
 derived guidelines for the design of future gene expression studies in
 species without sequenced genomes.
description: Dmel_denovo_assembly_VelvetOases.faD. melanogaster de novo assembly constructed using Velvet OasesDros_denovo_assembly_Trinity.faD. melanogaster de novo assembly constructed using TrinityDrosophila guided assembliesGuided assemblies constructed from D. melanogaster RNA-seq reads guided by the genomes of each of the other 11 drosophila species, constructed using Velvet Columbusggv.tgzhuman-RNAseq_mapto_Gorilla-gorillaHuman RNA-seq reads aligned to the G. gorilla genomehuman-RNAseq_mapto_Pan-troglodytesHuman RNA-seq reads aligned to the P. troglodytes genomehuman-RNAseq_mapto_Pongo-abeliiHuman RNA-seq reads aligned to the P. abelii genomehuman-RNAseq_mapto_Macaca-mulattaHuman RNA-seq reads aligned to the M. mulatta genomehuman-RNAseq_mapto_total-humanHuman RNA-seq reads aligned to the human genomedmel_RNA-seq_mapto_dana_samD. melanogaster RNA-seq reads aligned to the D. ananassae genomedmel_RNA-seq_mapto_dere_samD. melanogaster RNA-seq reads aligned to the D. erecta genomedmel_RNA-seq_mapto_dgri_samD. melanogaster RNA-seq reads aligned to the D. grimshawi genomedmel_RNA-seq_mapto_dmel_samD. melanogaster RNA-seq reads aligned to the D. melanogaster genomedmel_RNA-seq_mapto_dmoj_samD. melanogaster RNA-seq reads aligned to the D. mojavensis genomedmel_RNA-seq_mapto_dper_samD. melanogaster RNA-seq reads aligned to the D. persimilis genomedmel_RNA-seq_mapto_dpse_samD. melanogaster RNA-seq reads aligned to the D. pseudoobscura genomedmel_RNA-seq_mapto_dsec_samD. melanogaster RNA-seq reads aligned to the D. sechellia genomedmel_RNA-seq_mapto_dsim_samD. melanogaster RNA-seq reads aligned to the D. simulans genomedmel_RNA-seq_mapto_dvir_samD. melanogaster RNA-seq reads aligned to the D. virilis genomedmel_RNA-seq_mapto_dwil_samD. melanogaster RNA-seq reads aligned to the D. willistoni genomedmel_RNA-seq_mapto_dyak_samD. melanogaster RNA-seq reads aligned to the D. yakuba genomedmel_RNA-seq_mapto_dyak.sam.out.gzFigure_data_filesData text files for all figures. README.txt contains details of content.