蔡珊珊臺灣大學：植物科學研究所時雨青2007-11-272018-07-062007-11-272018-07-062005http://ntur.lib.ntu.edu.tw//handle/246246/57957Six pale yellow-pigmented organisms with optimal growth temperature around 50 oC, were isolated from hot springs in Taiwan. These strains were motile with single polar flagellum, non-spore forming, alkaliphilic, gram-negative rods. The G+C contents (mol %) of these isolates were ranged from 68.7 to 73.3. Phylogenetic analysis of the 16S rDNA and physiological, biochemical studies showed that these isolated can be classified into two groups. Group Ⅰ including isolates NTU-1000, NTU-1555 and NTU-1310 showed 99.6 % to 99.8 % sequences similarity to Silanimonas lenta 25-4T. All these strains can hydrolyze starch. The pH range for growth was 7.0-12.0, with an optimal at pH 9.0. Strains NTU-1000, NTU-1310, NTU-1555 and S. lenta 25-4T can grow in nutrient broth containing 1 % NaCl. Fructose was utilized by strains of Group Ⅰ and Silanimonas lenta 25-4T. NTU-1000, NTU-1310, NTU-1555 and S. lenta 25-4T produced the following enzymes: alkaline phosphatase; esterase(C4); esteraselipase(C8); lipase(C14); leucine arylamidase; trypsin; a-chymotypsin; acid phosphatase; Naphthol-AS-BI-phosphohydrolase. Group Ⅱ including isolates NTU-1133, NTU-1197 and NTU-1358 showed 94.8 % to 95.0 % sequences similarity to S. lenta 25-4T. Different from S. lenta 25-4T that hydrolysis of starch was not observed by these strains. Isolates NTU-1133 and NTU-1197 could grow in nutrient broth containing 2 % NaCl, however NTU-1358 and S. lenta 25-4T could grow at salt concentrations up to 3 %. Strain NTU-1133 and S. lenta 25-4T could assimilate cellobiose and D-glucose, however NTU-1197, NTU-1358 could not. Strain NTU-1197 could utilize mannitol; strain NTU-1133 could utilize raffinose and trehalose, but S. lenta 25-4T could not use these carbohydrates. Strains NTU-1133, NTU-1197, NTU-1358 and S. lenta 25-4T produced the following enzymes: alkaline phosphatase; esterase (C4); esteraselipase (C8); leucine arylamidas; trypsin; a-chymotypsin; acid phosphatase; Naphthol-AS-BI-phosphohydrolase. Differences in the characteristics of the strains were as follows: valine arylamidase and cystine arylamidase were positive in NTU-1133, NTU-1197, NTU1358 and a-glucosidase in NTU-1358. The major cellular fatty acids of NTU-1133, NTU-1197 and NTU1358 were 15:0 iso, 16:0 iso and 17:0 iso. 16:0 fatty acids were detected in the strains of Group Ⅱ but not in S. lenta 25-4T. Based on the DNA-DNA hybridization (low than 70 %) and the divergence in 16S rDNA (higher than 3 %). These three isolates (NTU-1133, NTU-1197, NTU1358) could be considered as new species of Silanimonas.中文摘要Ⅰ 英文摘要Ⅲ 縮寫對照表Ⅴ 表目錄Ⅵ 圖目錄Ⅶ 第一章 前言 1. 嗜熱性微生物 1 2. 嗜熱性微生物具熱穩定酵素 4 3. 嗜熱性微生物之研究 5 4. 嗜熱性蛋白細菌 6 第二章 材料方法 1. 實驗材料 10 2. 實驗方法 2.1 樣本採集 12 2.2 嗜熱性菌株分離、純化 13 2.3 菌種形態鑑定 13 2.4 細菌生理鑑定 15 2.5 脂肪酸組成分析 19 2.6 DNA G+C 含量之分析 20 2.7 16S rDNA片段序列分析 21 2.8 DNA-DNA 雜合反應 26 2.9 嗜熱菌的保存 30 2.10 資料分析 31 第三章 結果 1. 菌種分離、純化 33 2. 菌種形態鑑定 33 3. 細菌生理鑑定 35 4. 脂肪酸組成分析 39 5. DNA G+C 含量分析 39 6. 16S rDNA 序列分析 40 7. DNA-DNA 雜合反應 41 8. 16S rDNA 序列系統演化之分析 41 第四章 討論 1. 嗜熱性微生物研究之困難處 42 2. 菌種之分離 42 3. 嗜熱性微生物耐熱特性 44 4. 台灣本土新穎菌種之發現 45 5. 蛋白細菌之研究與應用 47 6. 總結 48 參考文獻 50 圖表 61 附錄一、10 X Castenholz basal salt solution 79 附錄二、菌種鑑定技術及基本流程圖 80 附錄三、菌種保存流程圖 81 附錄四、API-ZYM 酵素套件呈色對照表 82 附錄五、DNA 雜合反應之藥品、配方 83 附錄六、分離菌株之16S rDNA 序列 85734647 bytesapplication/pdfen-US嗜熱菌蛋白細菌thermophilicproteobacteriaotherhttp://ntur.lib.ntu.edu.tw/bitstream/246246/57957/1/ntu-94-R92b42002-1.pdf