The Nitrogen Fixation package provides the only safe and inexpensive means to measure nitrogenase activity in undergraduate laboratories. Traditionally, "N2 Fixation" has been measured by the acetylene reduction assay, which requires the use of a gas chromatograph to measure the reduction of acetylene to ethylene by N2 fixing organisms. Gas chromatographs are beyond the budget of most undergraduate courses, and the 10% acetylene required for the assay is explosive. In addition to these excellent reasons for avoiding the assay, it has been shown recently that the acetylene reduction assay inhibits the nitrogenase reaction!
Like photosynthesis, N2 Fixation is a fundamental biological process essential to global nutrient cycling, and as such it should be included in every undergraduate laboratory course in biology. To make this possible Qubit Systems has developed a Nitrogen Fixation Package that takes advantage of a natural product of the nitrogenase reaction (hydrogen) to measure nitrogenase activity. Also, unlike the acetylene reduction assay, the measurement of H2 allows N2 fixation rate to be calculated (rather than just total nitrogenase activity) and allows all measurements to be made in real time. In addition, the method is completely safe and very inexpensive.
The only limitation of the H2 assay for nitrogenase activity is that it can only be used with symbioses that lack the enzyme uptake hydrogenase (HUP). This enzyme oxidizes H2, and prevents its release from nodules. The package includes soybean seeds (Glycine max, cultivar Maple Arrow) and a peat-based inoculant of Bradyrhizobium japonicum strain 532C which, when used according to the plant cultivation section in this chapter, will produce H2-evolving legumes within a 28 day period. Other legumes may be used with the N2 Fixation package, but they should be inoculated with the correct (Brady-) Rhizobium strain lacking the HUP enzyme.
| Support and funding provided by: |
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Andrew W. Mellon Foundation |
Instrumentation manufactured by: |
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