Effect of Nitrate Induction, pH, Incubation Period and Chloramphenicol for Assayof Nitrite Reductase in Brassica napus L. Gupta Shilpa*, Atwal A K, Kumar Hitesh Department of Plant Breeding and Genetics, Punjab Agricultural University, Ludhiana-141 004, India *Author for correspondence: Email: shilpagupta09@gmail.com
Online published on 16 January, 2015. Abstract The present investigation commenced to standardize the optimum conditions for nitrite reductase enzyme after following some modifications like the effect of pH, prior induction with nitrate, additional incubation time and addition of chloramphenicol in its earlier available standard method. Three cultivars of Brassica napus L. viz. GSC-5, OCN-3 and GSL1 were raised under two different treatments i.e. without N and S application at the time of sowing (T0) and with 50 kg N/ha and 40 kg S/ha at the time of sowing (T1). Activity of the enzyme was found to be increased under anaerobic conditions and on addition of 20ìg of chloramphenicol. NIR activity was up shooted by prior incubation of tissue with nitrate (0.05M) for 12 hrs in both Canola (GSC-5 and OCN-3) and Non-Canola (GSL-1) type Brassica though higher in Canola species. Maximum NIR activity was observed under anaerobic conditions and at pH 4.5; followed by a linear decrease at pH 6.5, 7.5 and with no significant change at pH 8.5 under both treatments. Additional incubation of 40 min led to decrease in NIR activity under both aerobic and anaerobic conditions and under both treatments in three canola varieties. However there was no effect of anaerobic conditions and of prior induction with nitrate on nitrite uptake. Our study therefore reveals that optimum conditions for assay of NIR should be nitrate concentration of 0.05M, pH 4.5, 40 min. of incubation period and addition of 20ìg of chloramphenicol. Top Keywords Brassica napus, nitrite reductase, chloramphenicol, nitrate. Top |