Development of Methanoculleus-specific real-time quantitative PCR assay for assessing methanogen communities in anaerobic digestion.
J Appl Microbiol
; 116(6): 1474-81, 2014 Jun.
Article
em En
| MEDLINE
| ID: mdl-24521054
ABSTRACT
AIM:
To develop a Methanoculleus-specific real-time quantitative PCR (RT-qPCR) assay with high coverage and specificity for the analysis of methanogenic populations in anaerobic digestion. METHODS ANDRESULTS:
A Methanoculleus-specific primer/probe set for RT-qPCR was designed in this study based on all Methanoculleus 16S rRNA gene sequences in Ribosomal Database Project (RDP) according to TaqMan chemistry. The newly designed primer/probe set was shown to have high coverage and specificity by both in silico and experimental analyses. Amplification efficiency of the Methanoculleus-specific primer/probe set was determined to be ideal for RT-qPCR applications. Subsequent field testing on anaerobic digesters showed that results from RT-qPCR were consistent with those from clone library analysis, validating the accuracy of the RT-qPCR assay.CONCLUSIONS:
The Methanoculleus-specific RT-qPCR assay designed in this study can serve as a rapid and effective tool for the quantification of Methanoculleus populations in anaerobic digestion. SIGNIFICANCE AND IMPACT OF THE STUDY Methanoculleus populations represent important members of archaeal communities in methanogenic processes, necessitating the need to develop effective tools to monitor Methanoculleus population abundance. The RT-qPCR developed in this study provides an essential tool for the quantification of Methanoculleus populations in anaerobic digestion and for the understanding of the functions of these methanogens in anaerobic biotransformation.Palavras-chave
Texto completo:
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Sondas de DNA
/
Primers do DNA
/
Methanomicrobiaceae
/
Reação em Cadeia da Polimerase em Tempo Real
Idioma:
En
Revista:
J Appl Microbiol
Assunto da revista:
MICROBIOLOGIA
Ano de publicação:
2014
Tipo de documento:
Article
País de afiliação:
Estados Unidos