A second generation genetic linkage map of Japanese flounder (Paralichthys olivaceus).

BACKGROUND: Japanese flounder (Paralichthys olivaceus) is one of the most economically important marine species in Northeast Asia. Information on genetic markers associated with quantitative trait loci (QTL) can be used in breeding programs to identify and select individuals carrying desired traits. Commercial production of Japanese flounder could be increased by developing disease-resistant fish and improving commercially important traits. Previous maps have been constructed with AFLP markers and a limited number of microsatellite markers. In this study, improved genetic linkage maps are presented. In contrast with previous studies, these maps were built mainly with a large number of codominant markers so they can potentially be used to analyze different families and populations. RESULTS: Sex-specific genetic linkage maps were constructed for the Japanese flounder including a total of 1,375 markers [1,268 microsatellites, 105 single nucleotide polymorphisms (SNPs) and two genes]; 1,167 markers are linked to the male map and 1,067 markers are linked to the female map. The lengths of the male and female maps are 1,147.7 cM and 833.8 cM, respectively. Based on estimations of map lengths, the female and male maps covered 79 and 82% of the genome, respectively. Recombination ratio in the new maps revealed F:M of 1:0.7. All linkage groups in the maps presented large differences in the location of sex-specific recombination hot-spots. CONCLUSIONS: The improved genetic linkage maps are very useful for QTL analyses and marker-assisted selection (MAS) breeding programs for economically important traits in Japanese flounder. In addition, SNP flanking sequences were blasted against Tetraodon nigroviridis (puffer fish) and Danio rerio (zebrafish), and synteny analysis has been carried out. The ability to detect synteny among species or genera based on homology analysis of SNP flanking sequences may provide opportunities to complement initial QTL experiments with candidate gene approaches from homologous chromosomal locations identified in related model organisms.

Tiny RNAs associated with transcription start sites in animals.

It has been reported that relatively short RNAs of heterogeneous sizes are derived from sequences near the promoters of eukaryotic genes. In conjunction with the FANTOM4 project, we have identified tiny RNAs with a modal length of 18 nt that map within -60 to +120 nt of transcription start sites (TSSs) in human, chicken and Drosophila. These transcription initiation RNAs (tiRNAs) are derived from sequences on the same strand as the TSS and are preferentially associated with G+C-rich promoters. The 5' ends of tiRNAs show peak density 10-30 nt downstream of TSSs, indicating that they are processed. tiRNAs are generally, although not exclusively, associated with highly expressed transcripts and sites of RNA polymerase II binding. We suggest that tiRNAs may be a general feature of transcription in metazoa and possibly all eukaryotes.

Development of a DNA barcode tagging method for monitoring dynamic changes in gene expression by using an ultra high-throughput sequencer.

CAGE (cap analysis of gene expression) is a method for identifying transcription start sites by sequencing the first 20 or 21 nucleotides from the 5' end of capped transcripts, allowing genome-wide promoter analyses to be performed. The potential of the CAGE as a form of expression profiling was limited previously by sequencing technology and the labor-intensive protocol. Here we describe an improved CAGE method for use with a next generation sequencer. This modified method allows the identification of the RNA source of each CAGE tag within a pooled library by introducing DNA tags (barcodes). The method not only drastically improves the sequencing capacity, but also contributes to savings in both time and budget. Additionally, this pooled CAGE tag method enables the dynamic changes in promoter usage and gene expression to be monitored.