几丁质分解菌之研究.docVIP

幾丁質分解菌之研究 Study on Chitin-Degrading Bacteria 研究生：張淑貞 Chang, Shu-Chen 指導教授：吳明昌 Wu, Ming-Chang 江啟銘 Jiang, Chii-Ming 【摘要】 與傳統化學水解法相較，利用微生物直接將幾丁質轉化成的N-乙醯葡萄糖胺，具有高產率、低污染、以及具備專一性等優點；因此可大大提高轉化之產能。基於此，本研究架構一套平台以進行菌株分離，並予以鑑定和分類，以尋找嶄新且專一性佳、分解力強之幾丁質分解菌株；該成果除了在菌種分類學上具有貢獻，在實務上亦具有發展幾丁質寡醣量產技術之潛力。 本研究採集屏東縣不同區域的蝦類養殖水樣品，經純化後分離出可分解幾丁質之菌株。菌株經由革蘭氏染色後，僅保留革蘭氏陰性菌株；接著透過多相分析鑑定後，再以16S rDNA基因序列之演化分類學分析。經此程序共篩得三株新菌種，其中兩株編號為 cfT 與 c14T之分離菌株，經鑑定後為新屬菌株；而另一分離菌株編號為c1T經鑑定後為新種菌株。屬於 Betaproteobacteria綱，Neisseriales目，分別定名為「Chitinimonas taiwanensis gen. nov., sp. nov」，「Chitinilyticum aquatilis gen. nov., sp. nov」及「Chitinilyticum litopenae sp. nov」。此在分類學上有相當學術價值。 為了解新菌株之幾丁質分解能力與分解酶之特異性，本研究針對篩選之幾丁質分解菌所生產之代謝產物分析。結果顯示，此三菌株皆為具有高幾丁質分解能力之革蘭氏陰性桿菌；其中cfT以生產N-乙醯幾丁三醣為主，c14T以生產N-乙醯幾丁二醣為主，而c1T則以生產N-乙醯葡萄糖胺 和N-乙醯幾丁二醣為主。與已知之幾丁質分解菌株相較，三株新菌株皆具有更強之幾丁質分解能力。本研究成果可做為未來幾丁質酶與幾丁質寡醣生產之研究奠基。 關鍵字：幾丁質、演化分類、N-乙醯葡萄糖胺 【Abstract】 Compared with the traditional hydrolyze method for “chitin”- “N-acetyl-D-glucosamine (GlcNAc)” transferring process, the utilization of microorganism possesses many advantages, such as high yield, low pollution, and chitin-specificity. Therefore, this technique is helpful to raise the yield quantity of GlcNAc. In this study, we established a platform of bacteria isolation, identification and classification, which was able to find novel strains with good chitin-specificity and high chitin-degrading ability. The results in this study not only make good contribution to Taxonomy, but practically also have potentials to develop technique for mass productions. Samples were acquired by adopting aquatic water in various Pingtung regions; from which chitin-degrading strains were isolated through purification. Only Gram’s negative strains were preserved after Gram’s stain application. Subsequently these strains were characterized by a polyphasic approach, and then phylogenetic analyses based on 16S rRNA gene sequence were employed. Through the process three novel strains were obtained,