Design of PCR Primers for the <i>mcr</i>A Genes Detection of Methanogen Communities

Abstract

Methanogens play an important role to carbon cycling, catalyzing the production of methane and carbon dioxide, both potent green house gases, during organic matter degradation in anaerobic environments. Therefore, it is necessary to better understand microorganisms that produce natural gas. Indeed, methanogens are difficult to perform through culture based methods. In addition, the culture independent methods using the 16S rRNA gene also revealed some disadvantages. For these reasons, the culture independent molecular techniques using the specific catabolic genes such as methyl coenzyme M reductase (MCR) were studied. In this study, a primer set which can amplify specific fragments from a wide variety of mcrA gene was designed based on the homologous regions of 100 mcrA genes listed in the GenBank. PCR with the mcrA primers amplified DNA fragments of the expected size from all the six samples which obtained from biogas production reactors. In addition, denaturing gradient gel electrophoresis PCR analysis using our designed primers also revealed the diversity of mcrA gene in each sample. These results revealed that our primers were successfully to detect the mcrA genes and it is also helpful to know the diversity of mcrA genes in methanogen communities.