Optimization of the 15N2 incorporation and acetylene reduction methods for free-living nitrogen fixation

Smercina, Darian N.; Evans, Sarah E.; Friesen, Maren L.; Tiemann, Lisa K.

doi:10.1007/s11104-019-04307-3

Title: Optimization of the ¹⁵N₂ incorporation and acetylene reduction methods for free-living nitrogen fixation

Journal Article · 25 October 2019 · Plant and Soil

DOI: https://doi.org/10.1007/s11104-019-04307-3 · OSTI ID:1637663

^[1]; Evans, Sarah E. ^[2]; Friesen, Maren L. ^[3]; Tiemann, Lisa K. ^[1]

Michigan State Univ., East Lansing, MI (United States)
Michigan State Univ., Hickory Corner, MI (United States)
Washington State Univ., Pullman, WA (United States)

To optimize assay conditions of two common methods for measuring potential free-living nitrogen-fixation (FLNF), acetylene reduction assay (ARA) and ¹⁵N₂-incorporation (¹⁵N₂), for use with soil/rhizosphere samples. We tested the impact of different carbon (C) sources, oxygen concentrations (O₂), and incubation times on FLNF rates of two low-fertility Michigan soils via ARA and ¹⁵N₂.FLNF rates were greatest with addition of a C cocktail, at low O₂, and with 7-day incubations for both methods. FLNF via ARA was 1700x greater with a C cocktail versus glucose only and via ¹⁵N₂ was 17x greater with a C cocktail compared to other C sources and no-C controls. Specific O₂ optimum varied by method and site. A 7-day incubation was needed for the ARA, but a 3-day incubation was suitable for ¹⁵N₂. Lastly, we confirm previously identified issues with the ARA of acetylene-independent ethylene production/consumption resulting in potential FLNF measurement error of 1.3–52.3 μg N g^–1 day^–1. Here, we present an optimized method for measuring potential FLNF in soil/rhizosphere samples which will allow for consistent and comparable FLNF rate measurements. Researchers should account for C source, O₂, and incubation time when assessing FLNF and use the ARA method with caution.

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Research Organization:: Michigan State Univ., East Lansing, MI (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER)

Grant/Contract Number:: SC0018409; FC02-07ER64494; SC0014108

OSTI ID:: 1637663

Journal Information:: Plant and Soil, Vol. 445, Issue 1-2; ISSN 0032-079X

Publisher:: SpringerCopyright Statement

Country of Publication:: United States

Language:: English

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Cited by: 14 works

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