芽孢杆菌TS01的ARDRA评估和遗传分析

利用自主分离的芽孢杆菌菌株TS01和15种芽孢杆菌(地衣芽孢杆菌,枯草芽孢杆菌,短小芽孢杆菌,巨大芽孢杆菌,凝结芽孢杆菌,蜡状芽孢杆菌,迟缓芽孢杆菌,苏云金芽孢杆菌,嗜热脂肪芽孢杆菌,解淀粉芽孢杆菌,环状芽孢杆菌,球形芽孢杆菌,侧孢短芽孢杆菌,多粘类芽孢杆菌,泛酸枝芽孢杆菌)模式菌种进行ARDRA分析。采用16S rDNA通用引物16S-27和16S-1525进行PCR扩增，16S rDNA扩增片段经六种限制性酶（Alu I、Taq I、Mse I、Bst UI、Hha I和Tsp509 I）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和地衣芽孢杆菌处于同一分支，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16S rDNA序列分析结果一致，TS01是一株地衣芽孢杆菌菌株，从而证明ARDRA技术在菌种水平上对芽孢杆菌TS01进行鉴别具有可靠性。

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

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.megaterium,B.coagulans,B.cereus,B.lentus,B.thuringiensis,B.thermoaerophilus,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 Wag 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.