DNA Stable Isotope Probing

Abstract

The major obstacle for 15N-based DNA stable isotope probing (DNA-SIP) was insufficient separation of 15N-labeled and unlabeled DNA due to the lower nitrogen content in DNA compared with its carbon content. Extracted DNA can be loaded onto a cesium chloride (CsCl) gradient for isopycnic centrifugation and separation of labeled “heavy” DNA from unlabeled background DNA (“light” DNA). DNA-SIP experiments need to be implemented carefully in order to maximize achievable information and to avoid misinterpretation of resulting data. Using DNA-SIP, a study demonstrated the possibility that Candidate phylum TM7 is involved in toluene degradation in soils. Methanol, formaldehyde, and ammonium are the final products of the RDX degradation pathway. It is obvious that microorganisms that only use nitro-nitrogen in the ring-labeled 15N3-RDX would not be seen in this study. In a study, the authors used DNA-SIP to analyze the active methanotroph population in a peatland from the United Kingdom. By using DNA-SIP, the authors demonstrated that Methylocellaand Methylocystisare probably the most active methane utilizers in this peatland. DNA-SIP can be used in combination with metagenomics in a focused way to investigate the function of a subpopulation of environmental microorganisms. It is predicted that this approach will be adopted by more researchers in the near future. The 16S rRNA gene sequences retrieved from the SIP experiments were used to design specific probes targeting 16S rRNA gene of Acidovorax spp. and Pseudomonas spp.