Application of target capture sequencing of exons and conserved non-coding sequences to 20 inbred rat strains

AbstractWe report sequence data obtained by our recently devised target capture method TargetEC applied to 20 inbred rat strains. This method encompasses not only all annotated exons but also highly conserved non-coding sequences shared among vertebrates. The total length of the target regions covers 146.8Mb. On an average, we obtained 31.7× depth of target coverage and identified 154,330 SNVs and 24,368 INDELs for each strain. This corresponds to 470,037 unique SNVs and 68,652 unique INDELs among the 20 strains. The sequence data can be accessed at DDBJ/EMBL/GenBank under accession number PRJDB4648, and the identified variants have been deposited at http://bioinfo.sls.kyushu-u.ac.jp/rat_target_capture/20_strains.vcf.gz

A deletion in the intergenic region upstream of Ednrb causes head spot in the rat strain KFRS4/Kyo

[Background]Head spot is one of the phenotypes identified in the KFRS4/Kyo rat strain. Although previous linkage analysis suggested that Ednrb, which is frequently involved in coat color variations in various animals, could be the gene responsible for this phenotype, no mutations have been identified in its coding region. [Results]To identify mutations causative of this phenotype in KFRS4/Kyo, we analyzed target capture sequencing data that we recently generated. Our target capture method has a unique feature, i.e., it covers not only exonic regions but also conserved non-coding sequences (CNSs) among vertebrates; therefore, it has the potential to detect regulatory mutations. We identified a deletion of approximately 50 kb in length approximately 50 kb upstream of Ednrb. A comparative analysis with the epigenomic data in the corresponding region in humans and mice showed that one of the CNSs might be an enhancer. Further comparison with Hi-C data, which provide information about chromosome conformation, indicated that the putative enhancer is spatially close to the promoter of Ednrb, suggesting that it acts as an enhancer of Ednrb. [Conclusions]These in silico data analyses strongly suggest that the identified deletion in the intergenic region upstream of Ednrb, which might contain a melanocyte-specific enhancer, is the mutation causative of the head spot phenotype in the KFRS4/Kyo rat strain

Additional file 2: Figure S1. of A deletion in the intergenic region upstream of Ednrb causes head spot in the rat strain KFRS4/Kyo

PCR experiments to confirm the deletion found in KFRS4/Kyo. (PDF 68 kb

Additional file 4: Figure S3. of A deletion in the intergenic region upstream of Ednrb causes head spot in the rat strain KFRS4/Kyo

Virtual 4C output for the region around Ednrb gene in mouse genome (mm9 assembly). (PDF 308 kb

Additional file 1: of Design and application of a target capture sequencing of exons and conserved non-coding sequences for the rat

Table S1. Summary statistics for SNV and INDEL in various depths. Figure S1. Sequence coverage of the target regions for each rat strain. Figure S2. Number of homozygous SNVs identified in WTC/Kyo and PVG/Seac strains for each genomic region. Figure S3. Proportion of the number of SNVs in terms of the each class of regions in the target, i.e., CDS, UTR, CNS, and other regions, for each rat strain. Figure S4. The relationship between the phastCons conservation score and SNV density for each rat strain. (PDF 215 kb

Additional file 3: Figure S2. of A deletion in the intergenic region upstream of Ednrb causes head spot in the rat strain KFRS4/Kyo

A KFRS4/Kyo-specific deletion of approximately 50 kb in length located approximately 50 kb upstream of Ednrb. (PDF 116 kb