The activity of nitrate reductase (NR) is generally presumed to be a sensitive efficiency parameter and a reliable criterion to select crop genotypes with the highest capacity to utilize fertilizer nitrogen. In our experiments 3-day-old dark-germinated seedlings were transferred to light (22,000 lux, 16 h) and grown on KNOP solution. NR activity was measured in vitro in 6 to 9-day-old seedlings. NR activity reached maximal values in 7-day-old plants and 4 hours after the onset of illumination. Genotype-related variability of NR activities was almost 3-fold in inbreds and less than 2-fold in 18 studied crosses and their parental inbred strains (Table 2). The enzyme activity of any cross under study never exceeded that of its parental inbred strain with the highest NR activity. Significant variability among the genotypes seems to suggest gene-controlled rates of NR accumulation.
We investigated further to what extent maize genotypes differed in their rates of NR degradation. The rate of NR degradation (inactivation) was estimated in vitro at 0 C and 27 C. Two classes of inbreds were easily distinguished by their NR stability values (t 1/2 either 2 to 6 h or more than 18 h at 0 C). NR stability in vitro was not related to neutral (pH 7.6) protease activity: less than 20% variation of the latter was found in the inbred collection. The NR stability and activity values were also not interrelated, therefore these two parameters appeared to be inherited independently. The hybrids usually produced stable NR except in labile x labile crosses.
To estimate the NR activities in situ as the net nitrate reduction in intact tissues we measured either the accumulation of protein nitrogen or its enrichment with 15N during three successive 24-h intervals in 6 to 10-day-old seedlings. Two estimates agree perfectly well (r 0.82). To increase variability of NR levels, the experiments were performed using several genotypes. In the first series of experiments (inbreds M14, WF9, Oh43, Oh45 and crosses WF9 x Oh43, WF9 x Oh45, M14 x Oh45) protein accumulation values (15N method) taken for the combined data were in highly significant correlation with the NR activity values (df 25, r 0.87). The extension of the sample size in the second series of experiments provided a means to analyze such correlation separately for individual genotypes (Table 3). Both in the individual strains (except M14 and its hybrid) and in the whole block (the combined data excluding M14) the NR activity in vitro (the NR potential was significantly in line with the protein nitrogen accumulation (net nitrogen reduction) in the shoots and the roots. The NR potential and net nitrogen reduction were linear functions of shoot and root growth of seedlings. Yet the in vitro/in situ ratio was 10-fold in the shoots and 2-3-fold in the roots. As nitrate levels in the seedlings were shown to be non-limiting, this excess of the NR potential was presumably due either to partial NR inactivation or NADH compartmentation in vivo.
Thus it appears that by measuring NR activities in vitro at the seedling stage we are able to select those genotypes that absorb and reduce more nitrate. Yet without field experiments we cannot relate these data to the final crop yield and protein production.
L. P. Voronova, A. A. Peshkova and E. E. Khavkin
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