芽胞杆菌TS01的扩增核糖体DNA限制性分析(ARDRA)评估和遗传分析  

Evaluation of Amplified Ribosomal DNA Restriction Analysis (ARDRA) and Genetic Analysis of Bacillus TS01

作　　者：王程亮[1,2] 佟建明[1] 张潞生[2] 高微微[3] 李安英[1,2] 

机构地区：[1]中国农业科学院北京畜牧兽医研究所动物营养学国家重点实验室,北京100193 [2]中国农业大学农学与生物技术学院,北京100193 [3]中国医学科学院北京协和医学院药用植物研究所,北京100193

出　　处：《农业生物技术学报》2009年第6期1108-1113,共6页Journal of Agricultural Biotechnology

基　　金：国家科技支撑计划项目(No.2006BAD12B05)资助

摘　　要：利用自主分离的芽胞杆菌菌株TS01和15种芽胞杆菌(地衣芽胞杆菌(Bacillu.licheniformis)、枯草芽胞杆菌(B.subtilis)、短小芽胞杆菌(B.pumilus)、巨大芽胞杆菌B(.megaterium)、凝结芽胞杆菌(B.coagulans)、蜡状芽胞杆菌(B.cereus)、迟缓芽胞杆菌(B.lentus)、苏云金芽胞杆菌(B.thuringiensis)、嗜热脂肪芽胞杆菌(B.thermoaerophilus)、解淀粉芽胞杆菌(B.amyloliquefaciens)、环状芽胞杆菌(B.circulans)、球形芽胞杆菌(B.sphaericus)、侧胞短芽胞杆菌(Brevibacillus laterosporus)、多粘类芽胞杆菌(Paenibacillus polymyxa)和泛酸枝芽胞杆菌(Virgibacillus pantothenticus))模式菌种进行扩增核糖体DNA限制性分析(ARDRA)。采用16SrDNA通用引物16S-27和16S-1525进行PCR扩增,16SrDNA扩增片段经6种限制性酶(AluⅠ、TaqⅠ、MseⅠ、BstUⅠ、HhaⅠ和Tsp509Ⅰ)酶切电泳,获得了TS01菌株的特征性ARDRA指纹图谱。ARDRA图谱通过GelcomparⅡ软件进行聚类分析(UPGMA),结果表明,菌株TS01和地衣芽胞杆菌处于同一分支,亲缘关系最近。ARDRA分析鉴定结果与实验室前期菌株TS01形态、生化鉴定和16SrDNA序列分析结果一致,TS01是1株地衣芽胞杆菌菌株,从而证明ARDRA技术在菌种水平上对芽胞杆菌TS01进行鉴别具有可靠性。Bacillus TS01 is a functional strain separated by authors＇ laboratory. TS01 strain and other 15 Bacillus type strains （B. licheniformis, B. subtilis, B. pumilus, B. megatcrium, B. coagulans, B. cereus, B. lentus, B. thuringiensis, B. thcrmoaerophilus, B. amyloliquefaciens, B. circulans, B. sphaericus, Brevibacillus laterosporus, Paenibacillus polymyxa and Virgibacillus pantothenticus ） were used to identify the differentiation by amplified ribosomal DNA restriction analysis （ARDRA）. 16S rDNA was amplified by the general primer pair 16S-27 and 16S-1525. The 16S rDNA PCR amplicon （about 1.5 kb） was digested by six restriction enzymes （Alu Ⅰ , Taq Ⅰ, Mse Ⅰ, Bst U Ⅰ , Hha Ⅰ and Tsp509 Ⅰ ）. And the characteristic ARDRA fingerprint of TS01 was obtained. The ARDRA patterns were analyzed by Gelcompar Ⅱ software using UPGMA method. And the homology tree based on ARDRA analysis showed that TS01 strain belonged to B. licheniformis, which was identical to the identification results of TS01 by using morphology, biochemistry and 16S rDNA sequence analysis. And the results showed that the established technique of ARDRA analysis has the dependability for the identification of TS01 strain on the specific level .

关 键 词：芽胞杆菌TS01 ARDRA 遗传鉴定 

分 类 号：S188[农业科学—农业基础科学]