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Application of nested polymerase chain reaction for the detection of as yet uncultured organisms of the class Actinobacteria in environmental samples
Author(s) -
Rheims Holger,
Stackebrandt Erko
Publication year - 1999
Publication title -
environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.954
H-Index - 188
eISSN - 1462-2920
pISSN - 1462-2912
DOI - 10.1046/j.1462-2920.1999.00013.x
Subject(s) - biology , actinobacteria , 16s ribosomal rna , polymerase chain reaction , phylogenetic tree , library , ribosomal dna , clone (java method) , nested polymerase chain reaction , mega , environmental dna , genetics , botany , dna , bacteria , gene , ecology , biodiversity , physics , astronomy
Different groups of as yet uncultured, phylogenetically deeply rooting organisms of the class Actinobacteria have been shown previously by analysis of 16S rDNA clone libraries to be present in different soil and marine environments of worldwide distribution. In order to monitor specifically the occurrence of two of the actinobacterial groups among bacterial DNA extracted from different environments, a nested polymerase chain reaction (PCR) method, aiming at the sensitive detection of 16S rDNA, was used. Analysis of a clone library generated from one of these 16S rDNA‐derived, group‐specific PCR products of DNA from a forest soil broadened the spectrum of putative taxa within the two phylogenetic groups. Application of this PCR system demonstrated the presence of these organisms in natural environments and in a sample of dried and moistened peat. Fluctuations in the detectability of the target groups in different samples from the same site strongly suggest that the taxon diversity of a 16S rDNA clone library must be regarded as a temporal genomic snapshot taken from this environmental community.