以次世代定序技術解析台灣常見三種養殖鰻魚 (日本鰻、鱸鰻、太平洋雙色鰻) 之轉錄體及探討其基因表現概況

Abstract

日本鳗 (Anguilla japonica) 為台灣重要的經濟養殖魚種。然而，由於養鳗產業完全依賴天然鳗苗之供給，因此鳗苗的資源榮枯，決定了產業發展的永續性。近三十年來，日本鳗鳗苗之產量長期趨勢有明顯下降之現象，尤以近幾年為甚，鳗苗捕獲量頻創新低，一尾鳗苗身價高達 200 元，導致成鳗市場價格飆升，困擾著整個東亞的鳗魚產業。因此，近年來各國遂興起了異種鳗的養殖風潮，由國外引進大量異種鳗苗，其中鱸鳗 (A. marmorata) 以及太平洋雙色鳗 (A. bicolor pacifica) 是目前業界發展之重點。台灣地區已確認有四種鳗魚，最常見者為日本鳗及鱸鳗，另兩種太平洋雙色鳗與呂宋鳗 (A. luzonensis) 為熱帶品種，在台灣數量較少。然而，對於這些產業重要的養殖魚種，我們對於其基礎生物學，並沒有完整的研究資料平台可供參考。 本計畫利用NGS-illunina HiSeq 2000 次世代定序技術，將日本鳗、鱸鳗及太平洋雙色鳗之鳗苗全身組織的轉錄體定序，並進行組裝(de novo Assembly)。各物種都得到約10,000,000條 reads，各組裝出20萬餘條重組序列，平均長度大於800 bps。有40%以上的重組序列含有蛋白質編碼的區域 (protein coding region)。同時，分析KEGG資料庫的比對結果顯示，約有37%以上的組裝序列能在 KEGG中找到對應資料，約可涵蓋大部份的 KEGG 代謝途徑。 對於消化酵素基因分析，鱸鰻與太平洋雙色鳗鳗苗有相近的模式，可能兩者具有相似的營養需求，尤其是對於蛋白質的消化酵素表現遠高於日本鰻，代表二者對蛋白質的需求高於日本鰻。對於骨骼肌形進行分析，在玻璃鳗時期，慢速肌肉(slow muscle) 可能是主要的骨骼肌，其基因表現在鱸鰻與太平洋雙色鰻中高於日本鰻，應該是造成此三種鰻魚棲地不同的因素之一，使日本鰻主要分布於河川下游，而鱸鰻與雙色鰻則分布於河川上游區域。最後，建立轉錄體資料庫(ANGEL)，提供鳗魚基礎研究所需之資料。研究成果所建置之網路資料庫，已開放全世界連結運用，可提升我國在水產研究上的實力與能見度。

The Japanese eel (Anguilla japonica) is a commercially important aquaculture species in Taiwan. Because the full dependence of the glass eel supply caught from the wild for aquaculture, the available glass eel resource thus determines the sustainability of the eel aquaculture industry. In the past decades, glass eel abundances have been declining significantly, especially in the recent years. One piece of glass eel even costs as high as 200 NT dollars. This makes eel aquaculture industry suffer difficult condition. Therefore, the culture of other eel species becomes more and more popular during recent years. Abundant glass eels are imported from other countries, especially for the A. marmorata and A. bicolor pacifica. There are a total of 4 endemic eel species found in Taiwan, with Japanese eel and A. marmorata being the most common ones. The other 2 tropical eel species, namely A. bicolor pacifica and A. luzonensis, are rare in Taiwan. In present, the basic biological information of these aquaculture eel species, however, are still poor. In this study, we obtained transcriptome of A. japonica, A. marmorata and A. bicolor pacifica by Illunina HiSeq 2000. In each species, over 100 million reads were assembled into more than 200 thousand contigs (ave. length>800 bps). Over 40 % of contigs contain protein coding region. By KEGG pathway analysis, over 37% of contigs were annotated and involved in most KEGG metabolism pathway. A. marmorata and A. bicolor pacifica have similar gene expression patterns of digestive enzymes and may have similar dietary requirment, especially; the expression level of enzymes of protein is higher than A. japonica. It indicated that the requirement of protein for A. marmorata and A. bicolor pacifica is more than for A. japonica. In skeletal muscle analysis, slow muscle paly a major role in glass eel stage, and the expression level of gene of slow muscle is higher in A. marmorata and A. bicolor pacifica than in A. japonica. It suggested that the different expression level among three eel species may result in the difference of habitat usage. Thus A. japonica major lives in downstream and both of A. marmorata and A. bicolor pacifica are live in upstream. Finally, we constructed ANGEL database website, it may offer valuable information on biological research for scientists. The website opens to the world for free access. This can greatly increase the visibility of our science power on aquatic research.