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PMID: 16328656 Published · ppublish English Comparative Study Journal Article Research Support, Non-U.S. Gov't

Development of temporal temperature gradient electrophoresis for characterising methanogen diversity.

Microbial ecology ·Vol. 50 ·No. 3 ·2005-10-00 ·Pages 327-36

Earl J, Pickup RW, Ritchie DA, Edwards C

Abstract

Temporal temperature gradient electrophoretic (TTGE) analysis of 16S rDNA sequences was optimized to monitor the methanogen population present in water and sediments of a small eutrophic lake, Priest Pot, in the English Lake district. The production of nonrepresentative TTGE profiles due to the generation of polymerase chain reaction (PCR) artifacts initially proved problematical. The use of a proofreading polymerase in the PCR was found to be essential and fully optimized protocols were established and tested to ensure confidence that the TTGE profiles truly reflected sequence diversity. TTGE analysis revealed the methanogen population to be less diverse in water than in sediment. The most genetic diversity was observed in TTGE profiles of sediment DNA isolated in winter and the least was in sediment DNA isolated in summer. DNA sequencing analysis of bands recovered from TTGE gels revealed the presence of two methanogen communities. One clustered with Methanosaeta species and the other with the Methanomicrobiales. Many sequences showed low DNA sequence similarity to known methanogens, suggesting that Priest Pot harbors previously undescribed methanogen species.

MeSH Terms
DNA Primers DNA, Archaeal/analysis,genetics Electrophoresis, Gel, Two-Dimensional/methods Environmental Monitoring Fresh Water/analysis,microbiology Geologic Sediments/analysis,microbiology Methane/analysis Methanomicrobiales/classification,genetics,isolation & purification Methanosarcinales/classification,genetics,isolation & purification Molecular Sequence Data Polyethylene Glycols Polymerase Chain Reaction RNA, Archaeal/genetics RNA, Ribosomal, 16S/genetics Seasons Species Specificity Temperature United Kingdom Water Microbiology
Chemicals
DNA Primers DNA, Archaeal RNA, Archaeal RNA, Ribosomal, 16S Polyethylene Glycols Methane
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Earl Julie
School of Biological Sciences, University of Liverpool, Liverpool, L69 7ZB, UK.
Pickup Roger W
Ritchie Donald A
Edwards Clive
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Article Info
Journal
Microbial ecology
Abbr.
Microb Ecol
ISSN
0095-3628
DOI
10.1007/s00248-005-0192-1
Published
2005-10-00
Epub
2005-00-24
Pages
327-36
Language
English
Region
United States
NLM ID
7500663
Subset
IM
Databases
GENBANK
AY251025, AY251026, AY251027, AY251028, AY251029, AY251030, AY391857, AY391858, AY391859, AY391860, AY391861, AY391862, AY391863, AY391864, AY391865, AY391866, AY391867, AY391868, AY391869, AY391870, AY391871, AY391872, AY391873, AY391874, AY395082, AY395083, AY395084
External Links
PubMed source DOI: 10.1007/s00248-005-0192-1 View journal details
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