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研究生:劉雅芬
研究生(外文):YaFenLiu
論文名稱:以脈衝電泳法估算洛德乳酸桿菌的染色體大小並分析台灣雞隻腸道內此菌的分佈
論文名稱(外文):Genome size estimation of Lactobacillus reuteri DSM20016 and distribution analysis of L. reuteri strains isolated from gastrointestinal tract of chickens in Taiwan by pulsed-field gel electrophoresis
指導教授:張登欽張登欽引用關係
指導教授(外文):Tung-Ching Chung
學位類別:碩士
校院名稱:國立中興大學
系所名稱:獸醫微生物學研究所
學門:獸醫學門
學類:獸醫學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
論文頁數:72
中文關鍵詞:脈衝電泳法洛德乳酸桿菌染色體大小
外文關鍵詞:pulsed-field gel electrophoresisLactobacillus reuteriGenome size
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洛德因乳酸桿菌(Lactobacillus reuteri;LR)可分泌廣效性抗菌物質-洛德因(reuterin),為動物腸道中極為強勢(dominant)的有益菌種,LR的標準菌株DSM20016為Lerche及Reuter(1976年)自人糞便中分離到。本實驗以具高度區分(resolution)菌株能力之脈衝電泳(pulsed field gel electrophoresis;PFGE)分析法,先對DSM20016菌株進行染色體的大小(size)估算,經以NotI、SgfI及FseI三種酵素切割電泳後所產生之核酸片段總和,得知其平均值為2.1Mb。接著,本實驗以PFGE分析法,研究本省雞隻腸道中LR菌株的分佈情形,結果得知:(1)由各種雞種(土雞、肉雞、蛋雞)分離得之72株LR菌,可被本實驗所發展出之PFGE分析條件,分型出56種PFGE型,其分辨率(descriminative rate)高達78,0﹪,足見本省LR菌的多型性(clonal polymorphysm),(2)同一雞場之雞隻間,可採集到具相同PFGE型的LR菌(Type:L3、L8、L14、M1、E5、E13、E21),(3)不同場之雞隻間,亦可存在著相同PFGE型的LR菌(Type:M2、M4、E8),(4)自雞隻消化道中五個部位(嗉曩、腺胃、小腸、盲腸、結直腸)所篩選之LR菌,顯示L14、M1及E21三種PFGE型的LR菌,可在雞隻消化道的兩個部位以上同時被發現,此結果不但知道各部位間有不同PFGE型的LR菌存在,且首度證實同一種PFGE型的LR菌(Type L14),可同時存在於雞腸道之五個部位。
Lactobacillus reuteri, being able to secrete a strong microbiciidal substance termed reuterin, has been found as a dominant bacteria species in the gastrointestinal (GI) tract of animals. The type strain Lactobacillus reuteri DSM20016 was initially isolated from human feces by Lerche and Reuter in 1976. In this study, the pulsed-field gel electrophoresis (PFGE) technique was employed to estimate the genome size of Lactobacillus reuteri DSM20016. Through summation of the total DNA fragments generated by the restriction of L. reuteri chromosome with SgfI, NotI and FseI, respectively, the genome size was estimated as about 2.1Mb. Subsequently, the PFGE technique was used to study the distribution of L. reuteri strains in GI tract of chickens. Several important results were obtained as follows: (1) a total of 56 SgfI-PFGE types were identified from 72 strains of L. reuteri isolated from versatile chickens including native, broiler, and layer breeds, indicating a clonal polymorphysm of this species in Taiwan´s chicken, (2) some SgfI-PFGE types, including L3, L8, L14, M1, E5, E13, and E21 were found to exist in many chickens of a the same farm, simultaneously, (3) some SgfI-PFGE types, including M2, M4, and E8 were also found to exist in some chickens of different farms, (4) among the 5 GI portions investigated in this study, some SgfI-PFGE types such as L14, M1, and E21 were found to be capble of existing on more 2 GI portions, especially of the type L14 were isolated from the total 5 GI portions, simultaneously. This is the first evidence indcating the co-existance of a single PFGE type on 5 portions of GI tract.
目 錄
中文摘要
英文摘要
第一章 前言
第二章 文獻探討
第一節 乳酸菌在分子生物學方面之研究
第二節 洛德因乳酸桿菌之特性
第三節 脈衝電場膠體電泳(pulsed field gel electrophoresis, PFGE)技術之發展
第四節 脈衝電場膠體電泳技術之原理及限制
第五節 脈衝電場電泳膠體解析效果之影響因子
第三章 材料與方法
第一節 菌株來源及保存
第二節 脈衝電場電泳圖譜
第三節 洛德因乳酸桿菌之參考株Lactobacillus reuteri type strain DSM20016基因體大小之估算
第四節 本省各種雞隻(土雞、肉雞、蛋雞)腸道內之洛德因乳酸桿菌的基因型態分析
第五節 雞隻腸道各部位存在之洛德因乳酸桿菌基因型差異分析
第六節 分析比較實驗室保存之乳酸桿菌屬ATCC標準菌的基因型態差異
第七節 菌株plasmid干擾分型的分析
第四章 結果
第一節 脈衝電場膠體電泳圖譜限制酵素選擇
第二節 洛德因乳酸桿菌參考株DSM20016的基因體大小估算
第三節 台灣各種雞隻(包括土雞、肉雞、蛋雞)腸道內洛德因乳酸桿菌基因型態的分析
第四節 雞隻腸道各部位存在之洛德因乳酸桿菌基因型差異分析
第五節 菌株plasmid干擾分型之探討
第五章 討論
第六章 參考文獻
第七章 圖表
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