Open AccessSearching for predominant soil bacteria: 16S rDNA cloning versus strain cultivation
Author(s) -
Felske Andreas,
Wolterink Arthur,
Lis Robert,
Vos Willem M.,
Akkermans Antoon D.L.
Publication year - 1999
Publication title -
fems microbiology ecology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.377
H-Index - 155
eISSN - 1574-6941
pISSN - 0168-6496
DOI - 10.1111/j.1574-6941.1999.tb00642.x
Subject(s) - biology , 16s ribosomal rna , library , ribosomal dna , temperature gradient gel electrophoresis , cloning (programming) , actinobacteria , bacteria , microbiology and biotechnology , genetics , clone (java method) , gene , phylogenetics , computer science , programming language
The predominant bacteria in Dutch grassland soils, as identified by direct DNA extraction, PCR amplification of 16S rDNA and subsequent cloning and sequencing, were compared to the most abundant culturable bacteria. The 16S rDNAs of the strains from a comprehensive cultivation campaign were compared to some of the predominant cloned sequences by temperature gradient gel electrophoresis (TGGE). Four ribotypes were selected that were found to be abundant in the clone library: two closely related Bacillus ‐like sequences, a representative from the Verrucomicrobiales cluster and an uncultured member of the Actinobacteria. Using a variety of cultivation approaches a total of 659 pure cultures were isolated. Initially, approximately 8% of all isolates matched any of these ribotypes by same migration speed of their 16S rDNA amplicons on TGGE. However, sequencing analysis of matching isolates indicated that their 16S rDNA sequences were clearly different from the cloned sequences representing the fingerprint bands. Comparing the cultivation approach and the molecular 16S rDNA analysis from the same soil sample, there was no correlation between the collection of cultured strains and the 16S rDNA clone library.