Article (Scientific journals)
Testing of primers for the study of cyanobacterial molecular diversity by DGGE
Boutte, C.; Grubisic, Stana; Balthasart, Pierre et al.
2006In Journal of Microbiological Methods, 65 (3), p. 542-550
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Keywords :
16S ribosomal RNA; denaturating gradient gel electrophoresis; cyanobacteria; molecular diversity
Abstract :
[en] Denaturing Gradient Gel electrophoresis (DGGE) is a PCR-based technique which is widely used in the study of microbial communities. Here, the use of the three specific 16S rRNA cyanobacterial specific primers CYA359F, CYA781R(a) and CYA781R(b) on the assessment of the molecular diversity of cyanobacterial communities is examined. Assignments of the reverse primers CYA781R(a) and CYA781R(b) with cyanobacterial strain sequences showed that the former preferentially targets filamentous cyanobacteria whereas the latter targets unicellular cyanobacteria. The influence of the GC clamp position on the forward Or on reverse primer and the use of the two reverse primers separately or in equimolar mixture were investigated. Three environmental samples were subjected to amplification with 6 combinations of primers. The 6 banding patterns as well as the sequences of the bands extracted were analysed and compared. In addition, to assess the effect of the position of the GC clamp, the melting profiles of the sequences of Aphanizomenon flos-aquae PMC9707 and Synechococcus sp. MH305 were determined, with the GC clamp in the 3' or 5' position. Results showed that the use of two separate amplifications allowed a more complete study of the molecular diversity of the cyanobacterial community investigated. Furthermore, similar richness and identical phylogenctic assignments of extracted bands were obtained irrespective of the positioning of the GC clamp. (c) 2005 Elsevier B.V. All rights reserved.
Disciplines :
Microbiology
Biochemistry, biophysics & molecular biology
Author, co-author :
Boutte, C.
Grubisic, Stana ;  Université de Liège - ULiège > Services administratifs généraux > Protection et hygiène du travail (SUPHT)
Balthasart, Pierre ;  Université de Liège - ULiège > Département des sciences et gestion de l'environnement > Département des sciences et gestion de l'environnement
Wilmotte, Annick  ;  Université de Liège - ULiège > Département des sciences de la vie > Enzymologie
Language :
English
Title :
Testing of primers for the study of cyanobacterial molecular diversity by DGGE
Publication date :
June 2006
Journal title :
Journal of Microbiological Methods
ISSN :
0167-7012
eISSN :
1872-8359
Publisher :
Elsevier Science Bv, Amsterdam, Netherlands
Volume :
65
Issue :
3
Pages :
542-550
Peer reviewed :
Peer Reviewed verified by ORBi
Name of the research project :
MIDI-CHIP
Funders :
UE - Union Européenne [BE]
Available on ORBi :
since 26 August 2009

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