Presented at the Joint European Stable Isotope Users Group Meeting, Vienna, 30 August–3 September, 2004.
Technical considerations for the use of 15N-DNA stable-isotope probing for functional microbial activity in soils†
Article first published online: 9 MAY 2005
Copyright © 2005 John Wiley & Sons, Ltd.
Rapid Communications in Mass Spectrometry
Volume 19, Issue 11, pages 1424–1428, 15 June 2005
How to Cite
Cadisch, G., Espana, M., Causey, R., Richter, M., Shaw, E., Morgan, J. A. W., Rahn, C. and Bending, G. D. (2005), Technical considerations for the use of 15N-DNA stable-isotope probing for functional microbial activity in soils. Rapid Commun. Mass Spectrom., 19: 1424–1428. doi: 10.1002/rcm.1908
- Issue published online: 11 MAY 2005
- Article first published online: 9 MAY 2005
- Manuscript Revised: 6 MAR 2005
- Manuscript Accepted: 6 MAR 2005
- Manuscript Received: 1 NOV 2004
Stable-isotope DNA probing is a culture-independent technique that may provide a link between function and phylogeny of active microorganisms. The technique has been used in association with 13C substrates while here we evaluate feasibility and limitations of 15N-DNA stable-isotope probing (SIP) using labelled and unlabelled pure microbial cultures or soil extracts. Our results showed that 15N-DNA probing is feasible for cultures as well as soil samples. Limitations of 15N-DNA-SIP are (a) the need for relatively large quantities of DNA to visualise bands (although molecular resolution is much higher) and (b) 15N-DNA enrichment needed to ideally be >50 at%; however, this requirement can be lowered to approx. 40 atom% 15N with pure cultures using a modified CsCl centrifugation method (140K g for 69 h). These advances in 15N-DNA-SIP methodology open new opportunities to trace active microbial populations utilising specific N substrates in situ. Copyright © 2005 John Wiley & Sons, Ltd.