The use of denaturing gradient gel electrophoresis with fully-degenerate pufM primers to monitor aerobic anoxygenic phototrophic assemblages

Natalya Yutin, Oded Béjà,Marcelino T. Suzuki

Limnol. Oceanogr. Methods 6:427-440 (2008) | DOI: 10.4319/lom.2008.6.427

ABSTRACT: Aerobic Anoxygenic Photosynthetic bacteria (AAnP) were recently proposed to be significant contributors to the global ocean carbon and energy cycles, but relatively little is known about the spatial and temporal distribution of AAnP populations. Here, we report on the development of denaturing gradient gel electrophoresis (DGGE) analyses to monitor naturally occurring marine AAnP populations using the pufM gene. We developed an approach using fully degenerate universal pufM primers, and tested the effect of different primers and PCR conditions on DGGE band resolution and migration. PufM-DGGE analyses of eastern Mediterranean environmental DNA samples show that AAnP composition was coherent among stations and seasons. Furthermore, samples from different depths of stratified water columns resulted in clearly different DGGE patterns, while very similar DGGE patterns at varying depths were observed under well-mixed conditions. These results demonstrate the potential use of DGGE with fully degenerate primers as a tool for rapid analyses of natural marine AAnP communities.