Reassessing PCR primers targeting nirS, nirK and nosZ genes for community surveys of denitrifying bacteria with DGGE.

Abstract:

:We re-evaluated PCR primers targeting nirS, nirK and nosZ genes for denaturing gradient gel electrophoresis as a tool to survey denitrifying community composition in environmental samples. New primers for both nirS and nosZ were combined with existing primers, while for nirK the previously published F1aCu:R3Cu set was chosen for denaturing electrophoresis. All three sets yielded amplicons smaller than 500 bp and amplified the correct fragment in all environmental samples. The denaturing gradient gel electrophoresis worked satisfactorily for nirK and nosZ, but not for nirS. This was probably due to the multiple melting domains in this particular nirS fragment. From the excised and sequenced bands, only sequences related to the target genes were detected and tree analysis showed that the selected primers acted as broad range primers for each of the three genes. By use of the new nirS primers it was demonstrated that agricultural soil harbours a substantial diversity of nirS denitrifiers.

journal_name

FEMS Microbiol Ecol

authors

Throbäck IN,Enwall K,Jarvis A,Hallin S

doi

10.1016/j.femsec.2004.04.011

subject

Has Abstract

pub_date

2004-09-01 00:00:00

pages

401-17

issue

3

eissn

0168-6496

issn

1574-6941

pii

FEM401

journal_volume

49

pub_type

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