An improved zebrafish transcriptome annotation for sensitive and comprehensive detection of cell type-specific genes

Gene expression levels were quantified using RSEM. Median ratio normalized expression values are shown for each replicate, along with adjusted p-value, and log₂ fold change. Data used to generate plots in Figure 1A,B, and incorporated into source data tables indicated below.

Gene symbol, along with matching Ensembl gene ID and NCBI ID, as well as differential annotation (i.e., identified as differentially expressed only in RefSeq, Ens95, or both) are indicated. Expression data are derived from Figure 1—source data 1. Data used to generate plots in Figure 1C–G.

This file includes lists of Ens95 (worksheet 1) and RefSeq (worksheet 2) genes indicating annotation as coding sequence (CDS) and whether there is an annotated stop codon and 3' UTR. Data from RNA-seq-based quantification for Ens95 genes missing a 3' UTR that is present in RefSeq is included for kdrl^pos (worksheet 3), pdgfrb^pos (worksheet 4), and Nr2f2^pos (worksheet 5) cells. These data were used to generate Table 2 and graphs in Figure 2A,B; Figure 2—figure supplement 2I.

IDs for representative Ens95, RefSeq, and V4.3 transcript ID, along with V4.3 gene symbols are shown with respective 3' UTR lengths (worksheet 1). Average median ratio normalized expression and log₂ fold change (pos/neg) values quantified with Ens95, RefSeq, and V4.3 annotations from kdrl^pos (worksheet 2), pdgfrb^pos (worksheet 3), and Nr2f2^pos (worksheet 4) RNA-seq for reference genes are included. Data directly used to generate Figure 2D–G, Figure 2—figure supplement 2C–H, Figure 3B–J and incorporated into source data as indicated below.

Output from DESeq2 analysis comparing Nr2f2^pos and Nr2f2^neg RNA-seq from gene expression levels quantified using RSEM with Ens95 (worksheet 1) or RefSeq (worksheet 2). Median ratio normalized expression values are shown for each sample, along with adjusted p-value, p-value, log₂ fold change, fold change, and log₁₀ adjusted p-value. Intersection of genesets identified as significantly enriched in Nr2f2^pos cells using Ens95 or RefSeq (worksheet 3).

Worksheet one is a list of Ens95 genes missing from RefSeq with Ensembl gene ID, matching ZFIN ID and biotype annotation. Worksheet two is a list of RefSeq genes missing from Ensembl with NCBI gene ID, matching ZFIN ID, and coding sequence annotation. Transcript level matching output from gffcompare is included using Ens95 (worksheet 3) or RefSeq (worksheet 4) as a reference. Worksheet five is a transcript level comparison of Ens95 and Ens99. In this case, all transcripts exhibit a complete intron/exon chain match (designated by a ‘=" in class code). Data used to generate Table 3.

Table includes Ens95 gene symbol, gene ID, and spurious transcript ID. Persistence of observed discrepancy in Ens99 is indicated, as is previous status of curation in ZFIN. All of these have been reported to ZFIN.

This table includes information regarding blastx hits against zebrafish and human proteins, matches with lincRNAs, number of exons per gene, and whether the novel locus was included in the V4.3 annotation.

Annotation notes are also included regarding the relative strength of coordinate-based incorporation of NCBI (Entrez) and Ens99 gene identifiers.

Gene expression levels were quantified using RSEM with the V4.3 annotation. Median ratio normalized expression values are shown for each sample, along with adjusted p-value and log₂ fold change. Matching Ensembl and NCBI gene IDs are included.

Gene expression levels were quantified using RSEM with the V4.3 annotation. Median ratio normalized expression values are shown for each replicate, along with adjusted p-value and log₂ fold change. Matching Ensembl and NCBI gene IDs are included.

Gene expression levels were quantified using RSEM with the V4.3 annotation. Median ratio normalized expression values are shown for each replicate, along with adjusted p-value and log₂ fold change. Matching Ensembl and NCBI gene IDs are included. Worksheet 2 – Nr2f2^pos-enriched genes with matched entries from reference gene set (Figure 2—source data 2) and associated 3' UTR lengths (Figure 2—source data 2).

The p-value, adjusted p-value, and average log₂ fold change refer to comparison of indicated cluster with all other clusters. Cutoffs are adjp <0.05, log₂ fold change >0.5. Pct.1 is proportion of cells within indicated cluster that express detectable levels of the indicated gene. Pct.2 is proportion of all other cells that express detectable levels of the indicated gene. Matching LL and Ensembl gene IDs are included.

p-value, adjusted p-value and average log₂ fold change refer to comparison of indicated cluster with all other clusters. Cutoffs are adjp <0.05, log2 fold change >0.5. Pct.1 is proportion of cells within indicated cluster that express detectable levels of the indicated gene. Pct.2 is proportion of all other cells that express detectable levels of the indicated gene. Matching LL and Ensembl gene IDs are included.

Worksheet one includes all cartilage genes identified using both Ens95 and V4.2 quantification, as indicated, with associated adjusted p-values and log₂ fold change (comparison of cartilage cells to all other clusters). 3' UTR length from matching reference gene (from Figure 2—source data 2) is indicated. Worksheet two includes matched bulk RNA-seq quantification from Ens95 and V4.3 annotations for cartilage-specific genes. Data used to generate Figure 5A–C, Figure 5—figure supplement 1A,B.

Worksheet one includes all epidermis genes identified using both Ens95 and V4.2 quantification, as indicated, with associated adjusted p-values and log2 fold change. 3' UTR length from matching reference gene (from Figure 2—source data 2) is indicated. Worksheet two includes matched bulk RNA-seq quantification from Ens95 and V4.3 annotations for epidermis-specific genes. Data used to generate Figure 5A–C, Figure 5—figure supplement 1A,B.

First column is gene symbol, second column is assigned class shown in the Venn diagram.