A functional genomics tool for the Pacific bluefin tuna: Development of a 44K oligonucleotide microarray from whole-genome sequencing data for global transcriptome analysis

AbstractBluefin tunas are one of the most important fishery resources worldwide. Because of high market values, bluefin tuna farming has been rapidly growing during recent years. At present, the most common form of the tuna farming is based on the stocking of wild-caught fish. Therefore, concerns have been raised about the negative impact of the tuna farming on wild stocks. Recently, the Pacific bluefin tuna (PBT), Thunnus orientalis, has succeeded in completing the reproduction cycle under aquaculture conditions, but production bottlenecks remain to be solved because of very little biological information on bluefin tunas. Functional genomics approaches promise to rapidly increase our knowledge on biological processes in the bluefin tuna. Here, we describe the development of the first 44K PBT oligonucleotide microarray (oligo-array), based on whole-genome shotgun (WGS) sequencing and large-scale expressed sequence tags (ESTs) data. In addition, we also introduce an initial 44K PBT oligo-array experiment using in vitro grown peripheral blood leukocytes (PBLs) stimulated with immunostimulants such as lipopolysaccharide (LPS: a cell wall component of Gram-negative bacteria) or polyinosinic:polycytidylic acid (poly I:C: a synthetic mimic of viral infection). This pilot 44K PBT oligo-array analysis successfully addressed distinct immune processes between LPS- and poly I:C- stimulated PBLs. Thus, we expect that this oligo-array will provide an excellent opportunity to analyze global gene expression profiles for a better understanding of diseases and stress, as well as for reproduction, development and influence of nutrition on tuna aquaculture production

Interaksi Masyarakat Keturunan Arab dengan Masyarakat Setempat di Pekalongan

Dalam penelitian ini penulis mengeksplorasi interaksi antara masyarakat keturunan Arab dengan masyarakat setempat di Kelurahan Klego Kota Pekalongan serta mengetahui faktor pendorong dan penghambat terjadinya interaksi antara masyarakat keturunan Arab dengan masyarakat setempat. Penelitian ini menggunakan metode kualitatif. Teknik pengumpulan data melalui metode wawancara, observasi, dan dokumentasi. Hasil penelitian menunjukan bahwa terjadi interaksi antara masyarakat keturunan Arab dengan masyarakat setempat dengan intensitas dan kegiatan kebudayaan tertentu. Faktor pendukung terjadinya interaksi adalah adanya perkawinan campuran, terutama pada masyarakat keturunan Arab non-sayyid, dengan masyarakat setempat serta adanya kerjasama dalam bidang perdagangan. Sedangkan faktor penghambat terjadinya proses interaksi adalah adanya prasangka dan stereotip pada masyarakat keturunan Arab yang merasa masyarakat setempat kurang Islami, sebaliknya masyarakat setempat merasa masyarakat keturunan Arab itu sombong. Keturunan Arab yang tinggal di Kelurahan Klego terdiri dari golongan sayyid dan golongan non-sayyid. Keturunan Arab dari golongan non-sayyid sudah dapat berbaur dengan masyarakat setempat sedangkan keturunan Arab dari golongan sayyid belum berbaur dengan masyarakat non-Arab. Masyarakat keturunan Arab memiliki simbol-simbol seperti bahasa, pakaian, bangunan yang sangat mempengaruhi interaksi antara masyarakat keturunan Arab dengan masyarakat setempat. In this study, the author explores the interaction between people of Arab descent and the local people in the village of Klego Pekalongan city and also the factors that drive and inhibit the interaction between them. This study uses qualitative methods. The technique of collecting data are interviews, observation, and documentation. The results show that there is a pattern of interaction between people of Arab descent with the local people. Factors supporting the occurrence of interactions are the presence of mixed marriages, especially in the Arab non-sayyid descent, with the local community as well as the cooperation in the field of trade. While the factors inhibiting the interaction process is the existence of prejudice and stereotypes of people of Arab descent at a local community as less Islami. On the other hand, the local people feel that people of Arab descent are exclusive. The Arab descent living in the Village Klego consists of groups and classes of non-sayyid and sayyid. Arab descent from the class of non-sayyid are able to mingle with the local people, whereas Arab descent of sayyid cannot mingle with non-Arab communities. Society of Arab descent has symbols such as language, clothing, and building that strongly influence the interaction of people of Arab descent with the local community

Genetic parameters and quantitative trait loci analysis associated with body size and timing at metamorphosis into glass eels in captive-bred Japanese eels (Anguilla japonica).

The Japanese eel (Anguilla japonica) is among the most important aquaculture fish species in Eastern Asia. The present study aimed to identify the genetic parameters underlying body size and the timing at metamorphosis from leptocephali to glass eels in captive-bred Japanese eels, with the intent to foster sustainable development. Larvae from a partly factorial cross (14 sires × 11 dams) were reared until the point of metamorphosis into glass eels. In these organisms, we observed moderate heritability and mild genetic correlations among traits related to body size (h2 = 0.16-0.33) and timing at metamorphosis (h2 = 0.36-0.41). In an F1 full-sib family, quantitative trait loci (QTL) mapping for these traits identified one significant (genome-wide P < 0.05) and five suggestive QTLs (chromosome-wide P < 0.05). These results suggest that in the Japanese eel, metamorphic traits exhibit a polygenic genetic structure comprising many QTLs with small effects. In addition, we updated the genetic linkage map for the Japanese eel and integrated it with our newly constructed de novo genome assembly. The information and tools generated from this study will contribute to the development of freshwater eel genetics and genomics

Karyotype and FISH mapping probed with PCR amplicons of LG1-linked scaffolds in Japanese eel.

<p>Giemsa-stained karyotype of a wild-captured Japanese eel (A). Hybridization of rhodamine-labeled scaffold_127 amplicons (red signals) on a DAPI-stained metaphase spread (B). Hybridization of Alexa Fluor® 488-labeled scaffold_10233 (green signals) and rhodamine-labeled 12248 (red signals) amplicons on a DAPI-stained metaphase spread (C), and image of fluorescent signals only on the same metaphase spread (D). Arrowheads indicate hybridization signals. Scale bars represent 10 μm.</p

FISH mapping with putative LG1-linked scaffolds in Japanese eel.

<p>PCR amplicons from scaffold_12 (A), 135 and 297 (B), and 256 and 997 (C) are mapped to chromosome 5. The amplicons of scaffold_12 (A), 135 (B), and 256 (C) were labeled with rhodamine (red signals), and scaffold_297 (B) and 997 (C) were labeled with Alexa Fluor® 488 (green signals). Arrowheads indicate hybridization signals. Scale bars represent 10 μm.</p

Result of read mapping to the reference genome sequences of Japanese eel.

<p>Result of read mapping to the reference genome sequences of Japanese eel.</p

Identification of genetic linkage group 1-linked sequences in Japanese eel (<i>Anguilla japonica</i>) by single chromosome sorting and sequencing

<div><p>Japanese eel (<i>Anguilla japonica</i>) constitutes one of the most important food fish in Japan; accordingly, genome sequencing and linkage mapping have been conducted for the purpose of artificial cultivation. In the next stage, integration of genomic sequences within linkage groups (LG) is required to construct higher-resolution genetic markers for quantitative trait loci mapping and selective breeding of beneficial traits in farming. In order to identify LG1-linked scaffolds from the draft genome assembly (323,776 scaffolds) reported previously, we attempted to isolate chromosomes corresponding to LG1 by flow sorting and subsequent analyses. Initially, single chromosomes were randomly collected by chromosome sorting and subjected to whole-genome amplification (WGA). A total of 60 WGA samples were screened by PCR with primers for a known LG1-linked scaffold, and five positive WGA samples were sequenced by next-generation sequencing (NGS). Following reference mapping analysis of the NGS reads, four of the five WGA samples were found to be enriched by LG1-linked sequences. These samples were cytogenetically assigned to chromosome 5 by fluorescence <i>in situ</i> hybridization. Using blastn searches with 82,081 contigs constructed from the NGS reads of the four WGA samples as queries, 2323 scaffolds were identified as putative LG1-linked scaffolds from the draft genome assembly. The total length of the putative LG1-linked scaffolds was 99.0 Mb, comparable to the estimated DNA amounts of chromosome 5 (91.1 Mb). These results suggest that the methodology developed herein is applicable to isolate specific chromosome DNAs and integrate unanchored scaffold sequences onto a particular LG and chromosome even in teleost fishes, in which isolation of specific chromosomes by flow sorting is generally difficult owing to similar morphologies, sizes, and GC-contents among chromosomes in the genome. The putative LG1-linked scaffolds of Japanese eel contain a total of 6833 short tandem repeats which will be available for higher-resolution linkage mapping.</p></div

Chromosome painting with WGA products from single chromosomes.

<p>Chromosome painting probed with WGA products from sorted single chromosome samples EE2-sc-5sc (A), 6 (B), 8 (C), and 15 (D) on metaphase spreads from a Japanese eel cell line (EE2). The probes were labeled with rhodamine (red signals; A-C) or Alexa Fluor® 488 (green signals; D). Chromosome painting probed with a mixture of the four products on a metaphase spread of a wild-captured animal (E). Scale bars represent 10 μm.</p