Characterization of a de novo assembled transcriptome of the Common Blackbird (Turdus merula)

Abstract

BackgroundIn recent years, next generation high throughput sequencing technologies have proven to be useful tools for investigations concerning the genomics or transcriptomics also of non-model species. Consequently, ornithologists have adopted these technologies and the respective bioinformatics tools to survey the genomes and transcriptomes of a few avian non-model species. The Common Blackbird is one of the most common bird species living in European cities, which has successfully colonized urban areas and for which no reference genome or transcriptome is publicly available. However, to target questions like genome wide gene expression analysis, a reference genome or transcriptome is needed.MethodsTherefore, in this study two Common Blackbirds were sacrificed, their mRNA was isolated and analyzed by RNA-Seq to de novo assemble a transcriptome and characterize it. Illumina reads (125 bp paired-end) and a Velvet/Oases pipeline led to 162,158 transcripts. For the annotation (using Blast+), an unfiltered protein database was used. SNPs were identified using SAMtools and BCFtools. Furthermore, mRNA from three single tissues (brain, heart and liver) of the same two Common Blackbirds were sequenced by Illumina (75 bp single-end reads). The draft transcriptome and the three single tissues were compared by their BLAST hits with the package VennDiagram in R.ResultsFollowing the annotation against protein databases, we found evidence for 15,580 genes in the transcriptome (all well characterized hits after annotation). On 18% of the assembled transcripts, 144,742 SNPs were identified which are, consequently, 0.09% of all nucleotides in the assembled transcriptome. In the transcriptome and in the single tissues (brain, heart and liver), 10,182 shared genes were found.DiscussionUsing a next-generation technology and bioinformatics tools, we made a first step towards the genomic investigation of the Common Blackbird. The de novo assembled transcriptome is usable for downstream analyses such as differential gene expression analysis and SNP identification. This study shows the importance of the approach to sequence single tissues to understand functions of tissues, proteins and the phenotype.

De novo annotation of the transcriptome of the Northern Wheatear (Oenanthe oenanthe)

“…The transcriptome assembly of the wheatears yielded 21,982 contigs/isotigs, grouped into 17,539 isogroups. The putative number of coding genes identified in this communication, 15,640, is similar to other well-annotated birds’ transcriptomes, around 18,000 (see: Künstner et al, 2010 ; Balakrishnan et al, 2014 ; Finseth & Harrison, 2014 ; Koglin et al, 2017 ). We should mention that birds have suffered massive deletions of coding and non-coding regions in respect to reptilian sister clades and the rest of the vertebrates ( Huang et al, 2013 ; Lovell et al, 2014 ; Zhang et al, 2014 ) ( Fig.…”

Tendon-Derived Mesenchymal Stem Cells (TDSCs) as an In Vitro Model for Virological Studies in Wild Birds

“…Due to the absence of validated PCR primers or gene information on useful markers in blackbirds in the literature, they had to be predicted. For this purpose, the desired mRNA sequences were mapped from raw data from a transcriptomic analysis in blackbirds [ 30 ] using as reference the mRNAs of Swainson’s thrush [ 31 ], a species genetically close to the blackbird [ 32 ]. All bioinformatic analyses were performed using Geneious 10.2.3 software (Biomatters, Auckland, New Zealand).…”

“…From raw data of a Eurasian blackbird's transcriptomic analysis [30] we deduced the mRNA sequences from the positive TDSCs markers CD29, CD44, CD71, CD73, CD90, CD105, and the negative markers CD14, CD34, and CD45. From these mRNA sequences, we designed the primers presented in Table 1.…”

Avian transcriptomics: opportunities and challenges

“…At the time of this study, there was also no reference genome or transcriptome available for the Common Blackbird. This gap in our knowledge, however, is currently being closed as a new Common Blackbird de novo transcriptome annotation has recently been published based on 14 different tissues (Koglin et al 2017). …”

“…Ekblom et al 2014;Koglin et al 2017;Peterson et al 2012;Richardson et al 2017;Santure et al 2011;Srivastava et al 2012) (Box 1), sometimes in combination with the genome analyses of the same species (e.g. Chung et al 2015;Mueller et al 2015).…”