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Aquatic Microbial Ecology

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AME 54:211-216 (2009)  -  DOI:

Influence of primer mismatch and microdiversity on DGGE results: a case study with SAR11

Olga Sánchez1,*, Josep M. Gasol2,, Vanessa Balagué2, Ramon Massana2, Jordi Mas1, Carlos Pedrós-Alió2

1Departament de Genètica i Microbiologia, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain
2Departament de Biologia Marina i Oceanografia, Institut de Ciències del Mar CMIMA-CSIC, 08003 Barcelona, Spain

ABSTRACT: Although SAR11 is usually the dominant bacterial group in most marine ecosystems when analyzed with clone libraries and fluorescence in situ hybridization, it is often not retrieved in studies where denaturing gradient gel electrophoresis (DGGE) has been used. We analyzed the microdiversity of SAR11 in Blanes Bay (NW Mediterranean) and we suggest that the high evenness of multiple microdiverse phylotypes, none of which being particularly dominant, is the probable reason for this methodological discrepancy. We used seeding experiments in which different amounts of 2 SAR11-affiliated clones were mixed with DNA from an environmental sample obtained from the Blanes Bay Microbial Observatory. Two primer sets differing at 2 base positions produced DGGE images that varied in their SAR11 detection threshold concentration. Our results show that primer mismatches and/or the presence of faint bands due to microdiversity could explain why SAR11 is frequently not retrieved from DGGE gels.

KEY WORDS: DGGE · SAR11 · Microdiversity · Primers · Methodology

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Cite this article as: Sánchez O, Gasol JM, Balagué V, Massana R, Mas J, Pedrós-Alió C (2009) Influence of primer mismatch and microdiversity on DGGE results: a case study with SAR11. Aquat Microb Ecol 54:211-216.

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