qPCR and RT-qPCR Applied to Methane-cycling Archaea in the Marine Sediments of the White Oak River Estuary
Karen G. Lloyd
from: Applications of Molecular Microbiological Methods (Edited by: Torben L. Skovhus, Sean M. Caffrey and Casey R.J. Hubert). Caister Academic Press, U.K. (2014)
Quantitative PCR (qPCR) and reverse transcribed quantitative PCR (RT-qPCR) are powerful tools for quantifying the DNA and RNA, respectively, of specific groups of microorganisms in marine sediments. They can also be used to identify potential environmental functions of uncultured microorganisms by correlating microbial abundance and activity to geochemistry. An example of this usage is the present case study of the White Oak River estuary. Here uncultured anaerobic methane-oxidizing archaea (ANME) were shown to change in abundance (inferred from DNA quantified by qPCR) and activity (inferred from RNA quantified by RT-qPCR) in response to methane-oxidizing conditions, as well as methanogenic conditions. This non-culture-based method therefore raises the hypothesis that ANME archaea are capable of reversing their metabolism to methanogenesis. qPCR and RT-qPCR in marine sediments require careful checking to ensure maximal DNA and RNA extraction efficiencies, and minimial co-extraction of PCR inhibiting substances. If these pitfalls are avoided, qPCR and RT-qPCR can be used in other applications to develop hypotheses about the physiology of uncultured microorganisms in environmental samples read more ...