幾丁質酶 (EC 3.2.1.14) 為可分解幾丁質的一種醣類水解酶，先前實驗室已成功的在性喜以甲殼類為食的點帶石斑魚 (Epinephelus coioides) 的胃中選殖到兩個幾丁質酶基因，分別命名為幾丁質酶一及幾丁質酶二，並且發現此兩幾丁質酶可能與點帶石斑消化富含幾丁質之食物有關。本篇論文將就現有研究基礎進行更進一步的探討，本研究中將分為四大部分，分別針對 (一) 幾丁質酶基因結構分析、 (二) 幾丁質酶胺基酸序列演化樹建構、 (三) 幾丁質酶在魚苗發育階段的表現情形、 (四) 幾丁質酶基因表現的調控機制等進行探討。在研究中可發現到幾丁質酶一及幾丁酶二基因體去氧核醣核酸序列均是由11個外顯子及10個內插子所組成，但在序列全長上幾丁質酶二較幾丁質酶一長；在演化樹的建構分析上則可發現兩者均可歸類至脊椎動物幾丁質酶中的酸性幾丁質酶，但兩者在演化上則分屬於兩個不同支系；而在魚苗幾丁質酶基因表現分析實驗中則發現，當魚苗發育至第30~33天時，魚苗幾丁質酶基因表現量會有顯著性的增加；最後透過幾丁酶基因啟動子區域序列的選殖分析，則可發現到與脊椎動物胃蛋白酶基因表現調控有關的轉錄因子的鍵結區的存在，如：AML-1a、GATA-1、AP-1、IK-2等。從以上實驗可得知，點帶石斑魚的幾丁質酶一及二，可能均有與脊椎動物胃中消化酶相似的分子調控路徑，且兩者在魚苗發育中的表現情形也十分類似，但從演化樹的結果分析則可發現兩者分屬於不同支系，或許代表著兩者雖同在胃中表現且均與消化相關，但卻可能具有不同的生理意義。

Chitinase (EC 3.2.1.14) is a kind of glycosyl hydrolase which can hydrolysis chitin. Two chitinase genes (chitinase 1 and chitinase 2) have been cloned from the stomach of orange-spotted grouper (Epinephelus coioides). It is found that the chitinase was participating in the food digestions that contain the chitin in ingredient. In this study, four topics based on the previous data will be further discussing. The first is cloning the chitinase genomic DNA sequence. Second is to construct the phylogenetic tree of vertebrate chitinases. Third is to investigate the chitinase genes expression in the different development stages of the larvae. Fourth is to know the molecular mechanisms about the regulation of the chitinase genes expression. In the results, it could be found both of chitinase 1 and chtinase 2 are composed of 11 exons and 10 introns. Nevertheless, the whole length of chitinase 2 genomic DNA is much longer than chitinase 1. And in the phylogenetic tree, both of chitinase 1 and chitinase 2 are belong to the acidic chitinase of vertebrate chitinases, but they are in the different branches. Besides, by the analysis of chitinase genes expression in fish larvae, the genes are expressed significantly since 30~33 dpf (day post fertilization). Finally, through the cloning of chitinase genes promoter regions, both of them have the same transcription factors binding sites in the promoter region like the vertebrate pepsinogens. For example, the binding sites of AML-1a, GATA-1, AP-1 and IK-2. To make a summary of this study, the gene expression of chitinase 1 and chitinase 2 have the similar molecular regulatory way like the digestive enzyme in the stomach of vertebrate. In the fish larvae, the gene expression patterns of the two chitinases are similar. But in the phylogenetic tree, chitinase 1 and chitinase 2 are in the different groups. It may drop a hint, even though both chitinase 1 and chitinase 2 are expressed in stomach and have the same function about food digestion, but they may still have some different roles in physiology.

中華民國水產種苗協會(FBA) http://www.fish.org.tw/chinese/ Orange-spotted%20grouper.html
邵廣昭 台灣魚類資料庫 網路電子版 version 2009/1 http://fishdb.sinica.edu.tw, (2009-6-24)
曹鳳儒 (2007)，點帶石斑幾丁質酶純化、功能分析及基因選殖，國立成功大學生物科技研究所碩士論文
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