Optical bias with optical wedges in point sampling

Supplementary Tables and Figures. (PDF 454 kb

Additional file 4: of RNA secondary structure profiling in zebrafish reveals unique regulatory features

Number of read starts for every position covered in RNase V1 sample. (TXT 32051 kb

Additional file 9: of RNA secondary structure profiling in zebrafish reveals unique regulatory features

PARS scores of 54,083 transcripts and non-coding RNAs. (TXT 35418 kb

Additional file 5: of RNA secondary structure profiling in zebrafish reveals unique regulatory features

Number of read starts for every position covered in S1 nuclease sample. (TXT 45980 kb

Additional file 2: of RNA secondary structure profiling in zebrafish reveals unique regulatory features

Load score and percentage coverage of 54,083 transcripts. (TXT 3635 kb

Additional file 7: of RNA secondary structure profiling in zebrafish reveals unique regulatory features

Positions with ratio score more than one in V1 dataset. (TXT 15410 kb

Additional file 3: of RNA secondary structure profiling in zebrafish reveals unique regulatory features

Multi-conformation position counts in transcripts with overlapping peaks. (TXT 712 kb

Chamber Specific Gene Expression Landscape of the Zebrafish Heart

<div><p>The organization of structure and function of cardiac chambers in vertebrates is defined by chamber-specific distinct gene expression. This peculiarity and uniqueness of the genetic signatures demonstrates functional resolution attributed to the different chambers of the heart. Altered expression of the cardiac chamber genes can lead to individual chamber related dysfunctions and disease patho-physiologies. Information on transcriptional repertoire of cardiac compartments is important to understand the spectrum of chamber specific anomalies. We have carried out a genome wide transcriptome profiling study of the three cardiac chambers in the zebrafish heart using RNA sequencing. We have captured the gene expression patterns of 13,396 protein coding genes in the three cardiac chambers—atrium, ventricle and bulbus arteriosus. Of these, 7,260 known protein coding genes are highly expressed (≥10 FPKM) in the zebrafish heart. Thus, this study represents nearly an all-inclusive information on the zebrafish cardiac transcriptome. In this study, a total of 96 differentially expressed genes across the three cardiac chambers in zebrafish were identified. The atrium, ventricle and bulbus arteriosus displayed 20, 32 and 44 uniquely expressing genes respectively. We validated the expression of predicted chamber-restricted genes using independent semi-quantitative and qualitative experimental techniques. In addition, we identified 23 putative novel protein coding genes that are specifically restricted to the ventricle and not in the atrium or bulbus arteriosus. In our knowledge, these 23 novel genes have either not been investigated in detail or are sparsely studied. The transcriptome identified in this study includes 68 differentially expressing zebrafish cardiac chamber genes that have a human ortholog. We also carried out spatiotemporal gene expression profiling of the 96 differentially expressed genes throughout the three cardiac chambers in 11 developmental stages and 6 tissue types of zebrafish. We hypothesize that clustering the differentially expressed genes with both known and unknown functions will deliver detailed insights on fundamental gene networks that are important for the development and specification of the cardiac chambers. It is also postulated that this transcriptome atlas will help utilize zebrafish in a better way as a model for studying cardiac development and to explore functional role of gene networks in cardiac disease pathogenesis.</p></div

Data workflow and analysis summary.

<p>An overview of experimental and data analysis workflow adopted in the study for identification of known and putative novel cardiac chamber-restricted protein coding genes. (B) Venn diagram representing the total number of known protein coding genes (7,260) identified with expression level > = 10 FPKM across the three cardiac chambers. (C) A heat map representing expression pattern of 7,260 RefSeq protein coding genes with expression level > = 10 FPKM in the three cardiac chambers. The colour key represents transcripts in the range of 0 for transcripts with least expression to 15 for transcripts with maximum expression.</p

Differential expression of genes in the three cardiac chambers.

<p>The heat map represents expression profile of 96 cardiac chamber-restricted genes. The first column in the adjoining table lists the corresponding zebrafish gene names which are colour coded to highlight those with mutant phenotypes. Genes marked with green colour signify those which have been identified with heart related mutant phenotypes. The second column lists the human orthologs known for these genes. Genes marked with asterisks (*) represent those mentioned in the text (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0147823#sec012" target="_blank">Results</a> & <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0147823#sec023" target="_blank">Discussion</a>). Colour coding for the human orthologs highlights those which show disease associations. Orthologs marked with red colour are associated with cardiac disorders. The colour key represents transcripts in the range of 0 for transcripts with least expression to 15 for transcripts with maximum expression.</p