A simple method for measuring acetylene reduction of intact, nodulated black locust seedlings

A simple method is described for measuring acetylene (C(2)H(2)) reduction of nodulated black locust (Robinia pseudoacacia L.) seedlings. The C(2)H(2) reduction chamber consists of a standard plant pot sealed at one end with a stopper having holes for the seedling and a gas inlet port, and sealed at the other end with a stoppered collar with a gas port. The chamber temperature is regulated by circulating water at a controlled temperature through a copper coil surrounding the plant pot. A pump recirculates an approximately ten percent C(2)H(2) atmosphere through the system. Nine-week-old black locust seedlings, grown in sand culture and inoculated with Rhizobium, were used to obtain ethylene (C(2)H(4)) production curves. Ethylene production was linear for up to 60 minutes indicating that, for this particular symbiosis, (1) a simple closed system is adequate and (2) a short duration assay (10 minutes) will reliably estimate C(2)H(2) reduction over a longer period (up to at least 60 minutes). The water content of the soil medium had no influence on the C(2)H(2)/(C(2)H(2) + C(2)H(4)) ratio, which supports the suitability of C(2)H(2) as an internal standard for calculating C(2)H(2) reduction. This system has several desirable features. First, plant disturbance is minimized. Second, plants can be grown in inexpensive pots and no transplanting is necessary. Third, gas exchange and mixing are facilitated by a recirculating pump. Finally, root and nodule temperature can be controlled.