Identification of mixed bacterial DNA contamination in broad-range PCR amplification of 16S rDNA V1 and V3 variable regions by pyrosequencing of cloned amplicons
Identification of mixed bacterial DNA contamination in broad-range PCR amplification of 16S rDNA V1 and V3 variable regions by pyrosequencing of cloned amplicons
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DOI：
10.1016/s0378-1097(02)01190-4
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发表时间：
2003-02-14
影响因子：
2.1
通讯作者：
Jonasson, J
中科院分区：
文献类型：
--
作者：
Grahn, N;Olofsson, M;Jonasson, J
我们使用一种灵敏、快速的方法，结合细菌16S rDNA片段的宽范围PCR扩增和焦磷酸测序进行检测、鉴定和分型，我们发现了我们用于PCR的试剂中污染的细菌DNA。已鉴定的细菌为水媒细菌属假单胞菌属、寡养单胞菌属、黄单胞菌属、罗尔斯通氏菌属和芽孢杆菌属。我们的研究结果是一致的，与最近的报告污染的工业用水系统。鉴于这一结论，我们认为有必要提高对未经认证的广泛使用的分子生物学试剂（包括超纯水）可能污染的认识。由于基于序列的16S rDNA技术用于细菌分型和微生物群落表征的各种环境中，我们认为未来分子生物学试剂的认证（不含核酸）将是有利的。(C)2002年，欧洲微生物学会联合会。由Elsevier Science B.V.出版，版权所有。
Using a sensitive and rapid method combining broad-range PCR amplification of bacterial 16S rDNA fragments and pyrosequencing for detection, identification and typing, we have found contaminating bacterial DNA in our reagents used for PCR. Identified bacteria are the water-borne bacterial genera Pseudomonas, Stenotrophomonas, Xanthomonas, Ralstonia and Bacillus. Our results are in concordance with recent reports of contaminated industrial water systems. In light of this conclusion, we believe that there is a need for increased awareness of possible contamination in uncertified widely used molecular biology reagents, including ultra-pure water. Since sequence-based 16S rDNA techniques are used in a variety of settings for bacterial typing and the characterization of microbial communities, we feel that future certification of molecular biology reagents, as free of nucleic acids, would be advantageous. (C) 2002 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.