Nitrogen fixation, amino acid, and ureide associations in Chickpea.
Chickpea appears to be a warm-season crop or at least intermediate between warm and cool-season legumes on the basis of its [N.sub.2] fixation products. An earlier report suggested that chickpea exports ureides as the main N compound resulting from [N.sub.2] fixation, but the exact concentrations were not reported (Pate and Atkins, 1983). Later, amides were suggested to be the main nitrogenous compounds translocated out of chickpea nodules, with asparagine in the range of 40 to 70% of nodule xylem N and glutamine in the range of 5 to 45% nodule xylem N (Peoples et al., 1987). Further, ureides were not measured in their study.
Recent studies in soybean [Glycine max (L.) Merr.], a warm-season legume, implied that control of [N.sub.2] fixation during water deficit is mediated via cycling of metabolic products from the [N.sub.2] fixation process (Bacanamwo and Harper, 1997; Purcell et al., 2000). Sinclair and Serraj (1995) reported that warm-season species that accumulated high concentrations of ureides (>200 mmol [L.sup.-1] xylem sap) were more drought-sensitive than species with <50 mmol [L.sup.-1] xylem sap or no ureide. However, no comparable measurements of asparagine or glutamine have been taken. Previously, Serraj and Sinclair (1997) recorded ureides and total amino acids in xylem sap of eight soybean cultivars including Biloxi and Jackson, and the drought tolerant cultivars had the lowest concentration of ureides in the xylem and in petioles. In the soybean cultivars Biloxi and Jackson, the main xylem-transported amino acids were asparagine, glutamine, [gamma]-aminobutyric acid, and proline, but ureides were not measured (Serraj et al., 1998).
Past studies have suggested chickpea may be an amide transporting legume because chickpea is a cool season crop (Pate and Atkins, 1983). However, a recent study showed that ureide metabolism was also involved, because urea was found to be a product of ureidoglycolate degradation in different organs of chickpea (Munoz et al., 2001). In this study, the authors did not directly assay the enzyme step of ureide catabolism, which produces the urea.
To date, no conclusive studies have been performed in chickpea to determine the metabolic products of [N.sub.2] fixation in well-watered conditions or in drought, even though some research groups report amides and others report ureides. It is also not known if there are genotypic differences among chickpea cultivars in N metabolites and [N.sub.2] fixation under well-watered conditions. The objectives of our study were to determine the metabolites of [N.sub.2] fixation, namely free amino acids and ureides, in chickpea and to quantify the differences in [N.sub.2] fixation by [sup.15]N natural abundance among chickpea cultivars in the field under well-watered conditions.
MATERIALS AND METHODS
Field experiments were conducted in Saskatchewan, Canada, at one location in 2002 and two locations in 2003. The locations were Saskatoon (2002), Goodale (2003) and Saskatchewan Pulse Growers Land (SPG, 2003), and all locations were within a 30-km radius of Saskatoon, SK (52[degrees]09' N, 106[degrees]36' W). Soils at each location were Dark Brown Chernozems. Spring soil test values, average monthly precipitation and mean temperature during the summer at Saskatoon, 2002, and Kernen Farm Research Station, 2003 (within 15 km radius of Goodale and SPG locations) are presented in Table 1.
Cultivars Amit, CDC-Anna, CDC-Chico, CDC-Nika, and Myles were seeded in the field. These five chickpea cultivars were chosen on the basis of early maturity, leaf type (fern leaf), disease resistance, and high yield. Currently, these chickpea cultivars are best adapted to Dark Brown and Brown soils in Saskatchewan, Canada. Myles is widely grown in the northern USA, and the other cultivars have increasing production areas in North America. They were also the best five cultivars among eight previously tested in a growth chamber experiment, where selection was based on nodulation and their plant N requirement. The five cultivars and a reference crop were grown in each location in 2002 and 2003. Flax cv. CDC-Bethune was used as the non-fixing reference crop for the assessment of percentage N derived from the atmosphere (%Ndfa). Bremer and van Kessel (1990) suggested that the reference crop should have a similar rooting pattern and soil N uptake to the [N.sub.2] fixing plant. Flax, like chickpea, has a short tap root system with fibrous branches. Rooting depth for both crops extended to a depth of over lm depending on soil type. Plot size was 1.2 x 6.7 m with 0.3 m within and between row spacing. Commercial rhizobium inoculant was applied at the recommended rate (MicroBio RhizoGen Corp. Saskatoon, SK). Triple superphosphate (0-45-0) at the rate of 20 kg [ha.sup.-1] [P.sub.2][O.sub.5] was applied at seeding; no nitrogen fertilizer was applied. The residual N was less than 32 kg [ha.sup.-1] (Table 1). At each location, seeding was done by mid May and crops were grown until late vegetative stage before leaf sampling. The frost-free growing season at the sites was between 100 and 110 d. At Week 6 most of the cultivars at each location were in late vegetative growth and had begun to flower.
Six of the uppermost fully expanded leaves were chosen at random from the two center rows of each plot at each location at 6, 7, 8, 9, 10, and 11 wk after emergence. Final leaf tissue samples were taken at the end of August for each location and year. Leaflets were separated from the petiole, oven-dried (40[degrees]C for 2 d), finely ground, and analyzed for ureide concentration. Free amino acid concentrations in those sampled leaves were analyzed only at the Saskatoon location at 7, 9, and 11 wk after emergence. Biomass was removed from 1 [m.sup.2] of each plot at 6, 8, 10, and 12 wk after emergence for each location and biomass was oven-dried and finely ground. A sample of ground plant material was taken and analyzed for whole plant N content by combustion (LECO CNS 2000, St. Joseph, MI, USA). Plant N content was calculated by:
 Plant N content = Biomass(1 [m.sup.2]) x (%PlantN/100)
A 1-[m.sup.2] area of chickpea or reference crop at maturity was hand-harvested from each plot, dried, ball-milled, and analyzed for isotopic composition by the method described in Stevenson and van Kessel (1997) on a 20-20 Mass Spectrometer interfaced with an ANCA-GSL sample converter (Europa Scientific, Crewe, UK). Shoot materials were sampled at ground level. Any dropped leaves and the root system were not included in the sample. The proportion of N derived from the atmosphere via biological nitrogen fixation (%Ndfa) in chickpea shoot was calculated as reported by Rennie and Kemp (1984):
 %Ndfa = ([delta][sup.15][N.sub.flax] - [delta][sup.15][N.sub.chickpea]/[delta][sup.15][N.sub.flax] - C)100
where [delta][sup.15]N is:
 [delta][sup.15]N(%) = (atom%[N.sub.sample] - atom%[N.sub.atmosphere]/atom%[N.sub.atmosphere])1000
The value for C represents the [delta][sup.15]N value of chickpea grown in an N-free medium in a growth chamber under following conditions: 22[degrees]C during the day and 20[degrees]C during the night. The C value for the shoot of chickpea plants was 1.009. The atom percentage of [sup.15]N of the atmosphere was 0.3663 %, which was equal to a [delta][sup.15]N value of 0 (Mariotto, 1983). The amount of N fixed by each genotype was calculated by:
 [N.sub.2] fixed (kg [ha.sub.-1]) = (%Ndfa from shot/100) x (N yield from shot)
Seed yield was determined with a small plot combine for each location in early October.
A 30- to 35-mg sample of dry ground leaf sample from each plot was used to determine shoot ureide concentration using a modified colorimetric procedure (de Silva et al., 1996). Between 75 and 100 mg of homogenized oven dry leaf sample from each plot was used to extract free amino acids on the basis of a modified method of Leon-Guzman et al. (1997). One milliliter of C[H.sub.3]OH:CH[Cl.sub.3]:[H.sub.2]O (12:5:3) solution was added to 100 mg of leaf sample and shaken for 16 h on a reciprocating shaker. Then the mixture was centrifuged at 10 000 g for 5 min. A 100[micro]L aliquot of leaf extract was used to elute the free amino acids with the EZ: faast sample test kit (EZ: faast Phenomenex, Torrance, CA, USA). Free amino acids were analyzed on a gas chromatograph (6890 Series Gas Chromatograph Technologies, Agilent System, Wilmington, DE, USA) using a Zebron ZB-PAAC column (Phenomenex, Torrance, CA, USA). The EZ:faast method was developed for analysis of 40 aliphatic and aromatic amino acids (EZ: faast Phenomenex, Torrance, CA, USA). Mixtures of amino acid standards (20 nmol/100 [micro]L) were used for every 10 injections to quantify amino acid concentration. Norvaline was used as the internal standard and quantifications were performed by comparing sample peak areas to the standard's peak areas.
Experimental Design and Statistical Analysis
The experimental design was a randomized complete block design with four replicates, at three location-years. Sampling for the variables ureide concentration, amino acid concentration, and whole plant nitrogen content were taken from random samples within plots over time. Data were analyzed separately for each location-year. Ureide concentration, amino acid concentration, and whole plant nitrogen content were analyzed for each sampling time separately. Analysis of variance was done by the General Linear Model procedure (PROC GLM) of SAS version 8.2 (SAS Institute, 1999). Means were separated by Fisher's protected LSD at P < 0.05.
Soil Properties and Weather Data
The spring nutrient availability and weather data for each location are presented in Table 1. The soils of each location were clay loam. The pH ranged from 7.2 to 7.8 at 0- to 30-cm depth and no evidence of salinity was found. Spring available inorganic N concentration ranged from 22 to 32 kg [ha.sup.-1] in the 0- to 30-cm depth and the amount was higher in SPG location compared with Saskatoon and Goodale. Extractable P was low while extractable K was relatively high. About 30 to 40 g [kg.sup.-1] organic matter content was observed for each location in the 0- to 30-cm depth (Table 1). The Saskatoon location had above average precipitation from July to September, while Kernen had above average precipitation during April and July (Table 1). Generally, Goodale and SPG locations experienced dry conditions during summer compared with Saskatoon. The Saskatoon location had warm temperatures during June and July (2002), and the Kernen location had warm August temperatures in 2003.
Leaf Ureide Concentrations
Leaf ureide concentrations at Weeks 6 and 7 were low for all chickpea cultivars and ranged between 0.5 to 5 [micro]mol [g.sup.-1] (Table 2). Amit had a significantly higher ureide concentration compared with CDC-Chico at Week 6. Although Myles had a significantly higher leaf ureide concentration compared with CDC-Chico at Week 8, there were no significant differences among chickpea cultivars at Week 7. Myles showed significantly lower leaf ureide concentrations at Weeks 9, 10, and 11 compared with CDC-Chico. Cultivars were at late vegetative growth at Week 6 and flowering began at Week 8. There was a large increase in leaf ureide concentration of CDC-Chico after flowering. Furthermore, the amount of [N.sub.2] fixation by CDC-Chico was low compared with the other cultivars.
Mean Leaf Free Amino Acid Concentration
Alanine, asparagine, and glutamic acid were the major [N.sub.2] products resulting from [N.sub.2] fixation of chickpea and their concentrations were >70 [micro]mol [g.sup.-1] leaf dry weight (Table 3). Methionine, proline, serine, threonine, and valine were the second major [N.sub.2] products resulting from [N.sub.2] fixation. The remaining free amino acids were at concentrations <10 [micro]mol [g.sup.-1] of dry leaf tissue.
Alanine and asparagine concentrations over the sampling times were variable and differed depending on cultivar (Fig. 1). Alanine concentrations ranged between 135 [micro]mol [g.sup.-1] for CDC-Nika on Week 9 and 561 [micro]mol [g.sup.-1] for CDC-Nika on Week 11 when observing the variation across all cultivars from Weeks 7, 9, and 11. Asparagine concentrations ranged between 148 [micro]mol [g.sup.-1] for CDC-Nika on Week 9 and 333 [micro]mol [g.sup.-1] for Myles on Week 11 when observing the variation across all cultivars from Weeks 7, 9, and 11. Both alanine and asparagine concentrations decreased between Weeks 7 and 9 and then increased by Week 11 for an unknown reason (Table 3). Glutamic acid concentration increased at Week 9 to 121 [micro]mol [g.sup.-1] and then decreased at Week 11 to 79 [micro]mol [g.sup.-1] (Table 3). Methionine, proline, threonine, and valine increased over the sampling time but serine increased only at Week 9 to 35 [micro]mol [g.sup.-1] before decreasing to 31 [micro]mol [g.sup.-1] at Week 11 (Table 3). Aspartic acid, glutamine, histidine, isoleucine, leucine, lysine, and tryptophan increased over the sampling time (Table 3).
[FIGURE 1 OMITTED]
Overall, these results indicate that free amino acids are the major nitrogen metabolites compared with ureides. Most of the amino acids concentrations tended to increase at Week 11 except glutamic acid and glycine (Table 3). Most genotypic differences in free amino acid concentrations found in the leaves were observed at Week 9, which corresponded to late flowering and early pod formation.
Leaf Free Amino Acid Concentration Differences among the Cultivars
Cultivar differences in some free amino acids were observed at Weeks 9 and 11 (Fig. 1). At Week 7, no cultivar differences were observed except for isoleucine and serine (data not shown). Most cultivar differences in free amino acid concentrations in chickpea were observed at Week 9 (Fig. 1). CDC-Chico had a significantly higher alanine concentration compared with CDC-Nika at Week 9. Amit had a significantly higher asparagine concentration compared with CDC-Nika. Although CDC-Nika had lower alanine and asparagine concentrations at Week 9, its glutamic acid concentration was significantly higher compared with the other cultivars. CDC-Chico had a significantly lower asparagine concentration and a significantly higher glutamic acid concentration compared with Myles at Week 11.
Asparagine and alanine concentrations were higher in Myles at the beginning and at the end of the sampling period (Fig. 1). This may be due to Myles having an ability to maintain [N.sub.2] fixation products at a medium level throughout the growing season. However, Myles did have a significantly lower asparagine concentration compared with Amit at Week 9, indicating more N demand after flowering. Similarly, Myles had significantly lower concentrations of other amino acids such as glycine, lysine, phenylalanine, serine, and tryptophan at Week 9 compared with the other cultivars. The role of these amino acids in the metabolism of [N.sub.2] fixation products during drought is not yet known.
Whole Plant N Content, Seed Yield, and Percentage N Derived from Atmosphere
As anticipated, whole plant N content increased for all chickpea cultivars over the growing season; however, a significant cultivar effect was observed only at Weeks 6 and 10 (Table 4). At Week 6, CDC-Nika had a significantly higher plant N content compared with CDC-Anna and Myles. Amit had a significantly higher plant N content compared with Myles at Week 10. Seed yield of CDC-Anna from the three sites was significantly lower than the other chickpea cultivars (Table 4). Seed yield for CDC-Anna in 2002 was low at Saskatoon because of frequent rain, flood, and disease. The seed yield average over two locations in 2003 for CDC-Anna ranged between 1100 and 1500 kg ha-l, which was similar to the other chickpea cultivars. Although CDC-Anna had lower seed yield, %Ndfa was significantly higher than CDC-Chico and Amit. Myles, CDC-Nika and CDC-Anna were the highest [N.sub.2] fixing cultivars and CDC-Chico was the lowest. Myles did not have a significantly lower total N content, meaning that the moderate concentrations of ureides are not a result of lower [N.sub.2] fixation but likely a continued and steady metabolism.
Asparagine and alanine were the major shoot free amino acids found, implying these are candidates for metabolic products resulting from N fixation, but chickpea also produced ureides that accumulated in leaves to between 0.5 to 3.8 [micro]mol [g.sup.-1] dry weight. This is the first study reporting chickpea cultivar variation in free amino acids associated with [N.sub.2] fixation under field conditions and that chickpea also has a high concentration of alanine (up to 560 [micro]mol [g.sup.-1]).
Chickpea has [N.sub.2] metabolism that is comparable with the ureide exporting legumes soybean and cowpea (Hong and Copeland, 1990), so ureides would be expected as metabolic products. Similar to the Hong and Copeland (1990) study, Munoz et al. (2001) found catabolic ureide enzyme activities. They reported that the presence of the ureidoglycine aminohydrolase enzyme complex increased the production of ureidoglycolate in chickpea leaves. The presence of ureidoglycolate urealyase or ureidoglycine aminohydrolase further breaks ureidoglycolate into glyoxylate. The presence of ureidoglycolate urea-lyase activity demonstrates the existence of a urea-producing pathway for ureide catabolism in chickpea. This means chickpea has an ability to produce ureides as metabolites resulting from [N.sub.2] fixation. Results from Hong and Copeland (1990) and Munoz et al. (2001) fit our experimental results in that both asparagine and ureides can be major shoot metabolites resulting from [N.sub.2] fixation in chickpea. However, from our data, asparagine is the free amino acid found at the highest concentration, 213 to 290 [micro]mol [g.sup.-1] of dry leaf tissue (with a ratio of 2 moles N for every mole of asparagine), and allantoin and allantoic acid are at lower concentrations, with about 1 to 4 [micro]mol [g.sup.-1] of dry leaf tissue (with a ratio of 4 moles N for every mole of ureide). We found chickpea cultivars maintained ureide concentrations during the growing season, but poor [N.sub.2] fixing chickpea cultivars showed high ureides and low asparagine concentrations at the end of the growing season.
Under field conditions in Syria, Beck (1992) reported %Ndfa value for chickpea ranged from 0 to 80% and the average value for N2 fixation ranged from 19 to 24 kg N [ha.sup.-1] during a dry year. Carranca et al. (1999) reported %Ndfa in developing pods of chickpea ranged from 30 to 80% under field conditions with or without inoculants. Although, %Ndfa values from the straw and pod of chickpea were similar, Carranca et al. (1999) suggested that chickpea remobilized large amounts of N from vegetative parts to pods during reproductive stage. Typically under dryland conditions shoot N derived from [N.sub.2] fixation in chickpea represents to 40 kg N [ha.sup.-1] and this amount of [N.sub.2] fixation appears to be similar or marginally higher under irrigated conditions (Unkovich and Pate, 2000). Values of %Ndfa and [N.sub.2] fixed by chickpea in our experiment were similar to the results reported by Beck (1992) and Carranca et al. (1999). Amit and CDC-Chico had lower %Ndfa values compared with Myles. Myles and CDC Chico represented the range of nitrogen fixation (10-50%) seen in cultivars grown in western Canada. Possible reasons for low %Ndfa in Amit and CDC-Chico were the high ureide and glutamic acid concentrations, coupled with a low asparagine concentration at the end of the growing season. A labeling study has shown that accumulation of asparagine and ureides increases the pool of soluble N in faba bean (Vicia faba L.) that can cause a feedback inhibition effect on the nitrogenase activity (Oti-Boateng and Silsbury, 1993). CDC-Chico has an indeterminate growth habit and grows to a large size with many branches, and shows N deficiencies after flowering in both the field and growth chamber. Late season symptoms of N deficiency m such cultivars are consistent with a high ureide concentration after flowering, which may cause a feedback effect on [N.sub.2] fixation and which subsequently reduces asparagine concentration.
The cultivar Myles maintained ureides and amides at a moderate concentration for a longer time compared with the other tested chickpea cultivars in the field. Asparagine was the principle export product that was accumulated in young pea leaves under drought conditions (Ta et al., 1985). Asparagine synthesis occurs via a glutamine-dependent amidation of aspartate. Active transport mechanisms may contribute to low amino acid concentration in nodules. Active transport of amino acids can then create a greater diffusion gradient between symbiosome and cytosol that may cause a feed back inhibition on the nitrogenase activity (Ta et al., 1985).
This research was funded by NSERC and Saskatchewan Pulse Growers Association in Canada.
Abbreviations: Ec, electrical conductivity: %Ndfa, percentage nitrogen derived from the atmosphere; SPG, Saskatchewan Pulse Growers.
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Dil Thavarajah, Rosalind A. Ball, * and Jeff J. Schoenau
Dil Thavarajah and Rosalind A. Ball, Department of Plant Sciences, and Jeff J. Schoenau, Department of Soil Science, University of Saskatchewan, 51 Campus Dr., Saskatoon, Saskatchewan, Canada S7N5A8. * Corresponding author (email@example.com).
Table 1. Soil physicochemical properties, precipitation, and mean temperature for the 2002 and 2003 growing seasons at three locations near Saskatoon, SK. Soil physicochemical properties Electricity Location and year Texture pH conductivity N[O.sub.3]-N kg [ha. mS [cm.sup.-1] sup.-1] Saskatoon, 2002 Clay loam 7.2 0.2 22 Goodale, 2003 Clay loam 7.3 0.3 24 SPG, 2003 Clay loam 7.8 0.2 32 Monthly precipitation mm April May June July Saskatoon, 2002 10 2 52 70 Kernen, 2003 61 14 31 64 30-yr average Saskatoon 25 44 63 58 Kernen 24 44 61 57 Mean air temperature [degrees]C April May June July Saskatoon, 2002 -0.5 9 17 20 Kernen, 2003 5 12 16 18 30-yr average Saskatoon 6 12 16 19 Kernen 5 12 16 18 Soil physicochemical properties Organic Location and year P K matter kg [ha.sup.-1] g [kg.sup.-1] Saskatoon, 2002 46 970 35 Goodale, 2003 45 980 35 SPG, 2003 37 848 39 Monthly precipitation mm August September Total Saskatoon, 2002 75 49 258 Kernen, 2003 31 25 226 30-yr average Saskatoon 37 32 259 Kernen 35 33 254 Mean air temperature [degrees]C August September Saskatoon, 2002 17 11 Kernen, 2003 21 10 30-yr average Saskatoon 17 11 Kernen 18 11 Table 2. Leaf ureide concentration ([micro]mol [g.sup.-1] dry leaf tissue) for field-grown chickpea cultivars. Weeks after emergence Year Location Genotypes 6 7 8 Leaf ureide concentration [micro]mol [g.sup.-1]dry leaf tissue 2002 Saskatoon Amit 4.5a 3.0a 4.9ab ([dagger]) CDC-Anna 2.8ab 2.5a 5.9ab CDC-Chico 1.4b 4.7a 3.3b Myles 2.3ab 4.5a 6.7a CDC-Nika 3.1ab 4.7a 4.4ab P value 0.2 0.8 0.1 Standard error 0.87 1.78 0.86 2003 Goodale Amit 3.9a 1.1a 1.5a CDC-Anna 2.1a 2.5a 1.8a CDC-Chico 2.1a 1.7a 1.1a Myles 2.3a 2.4a 1.4a CDC-Nika 3.6a 1.8a 0.3a P value 0.5 0.6 0.6 Standard error 1.05 0.2 0.067 2003 SPG Amit 1.6a 0.5a 0.8a CDC-Anna 1.1a 0.9a 0.6a CDC-Chico 1.1a 0.7a 1.1a Myles 1.3a 1.1a 1.7a CDC-Nika 1.4a 1.1a 0.7a P value 0.3 0.3 0.2 Standard error 0.18 0.25 0.34 Weeks after emergence Year Location Genotypes 9 10 11 Leaf ureide concentration [micro]mol [g.sup.-1]dry leaf tissue 2002 Saskatoon Amit 4.7a 6.5a 5.4ab CDC-Anna 2.6b 3.1bc 6.8ab CDC-Chico 5.6a 5.3ab 8.9a Myles 2.0b 1.3c 4.8b CDC-Nika 2.5b 3.4bc 6.6ab P value 0.04 0.002 0.2 Standard error 0.59 0.73 1.19 2003 Goodale Amit 2.8ab 0.5b 0.5bc CDC-Anna 1.7abc 0.5b 0.9ab CDC-Chico 3.3a 1.4a 1.1a Myles 1.2bc 0.9ab 0.4c CDC-Nika 0.6c 0.5b 0.4c P value 0.07 0.01 0.02 Standard error 0.67 0.18 0.17 2003 SPG Amit 1.9a 3.6ab 4.3a CDC-Anna 1.8a 3.5ab 2.8a CDC-Chico 2.0a 6.2a 4.2a Myles 3.9a 2.3b 1.5a CDC-Nika 1.7a 4.3ab 3.9a P value 0.2 0.2 0.3 Standard error 0.76 1.1 1.06 ([dagger]) Comparisons made each week separately. Means within a column (week) followed by the same letter are not significantly different at P < 0.05. Table 3. Mean leaf free amino acid concentration during reproductive growth, averaged over five chickpea cultivars grown in the field, Saskatoon, 2002. Weeks after emergence Free amino acid 7 9 11 Mean free amino acid concentration [micro]mol [g.sup.-1] of dry leaf tissue Alanine 350.4 309.9 397.8 Asparagine 239.2 212.7 * 289.9 * Aspartic acid 1.4 2.0 4.8 * Glutamine 2.0 1.9 * 3.1 Glutamic acid 82.5 121.3 * 79.1 Glycine 3.1 4.3 * 3.1 Histidine 2.5 3.8 5.5 Hydroxyproline 30.8 25.3 * 14.9 * Isoleucine 5.2 5.5 6.1 Leucine 7.2 * 19.3 25.9 Lysine 0.5 0.7 * 1.3 Methionine 16.9 38.8 * 48.4 Phenylalanine 9.9 6.6 * 8.1 Proline 19.9 26.4 32.4 Serine 29.7 * 35.4 * 31.0 Threonine 13.1 18.7 * 26.8 Tryptophan 9.0 13.5 * 10.2 Valine 11.3 13.8 * 25.9 * The concentration of a specific amino acid for that specific week (7, 9, or 11) differed significantly among cultivars at P < 0.05. Table 4. Whole plant N, grain yield, percentage N derived from the atmosphere, and amount of [N.sub.2] fixation for field-grown chickpea cultivars. Weeks after emergence-whole plant N Year Location Genotypes 6 8 10 g N [m.sup.-2] 2002 Saskatoon Amit 3a 8a 11a ([dagger]) CDC-Anna 2a 6a 9.8ab CDC-Chico 3a 8a 10.3ab Myles 3a 6a 8.3b CDC-Nika 3a 8a 10.7ab P value 0.7 0.4 0.2 Standard error 0.39 1.97 1.96 2003 Goodale Amit 2.7abc 6a 9a CDC-Anna 2.5bc 6a 8a CDC-Chico 3.2ab 5a 12a Myles 2.4c 5a 10a CDC-Nika 3.4a 5a l0a P value 0.05 0.6 0.4 Standard error 0.23 0.79 1.42 2003 SPG Amit 2.3ab 4a 12a CDC-Anna 2.3ab 4a 12a CDC-Chico 2.0ab 4a 11a Myles 1.6b 4a 10a CDC-Nika 2.6a 5a 12a P value 0.2 0.5 0.4 Standard error 0.28 0.52 0.97 Weeks after emergence-whole plant N Grain Year Location Genotypes 12 yield g N [m.sup.-2] kg [ha.sup.-1] 2002 Saskatoon Amit 11a 797a CDC-Anna 13a 304b CDC-Chico 9a 853a Myles 10a 747a CDC-Nika l0a 862a P value 0.4 0.01 Standard error 1.0 110 2003 Goodale Amit 7a 1487a CDC-Anna 9a 1191a CDC-Chico 10a 1383a Myles 9a 1568a CDC-Nika 6a 1445a P value 0.5 0.5 Standard error 1.56 177 2003 SPG Amit 14a 3180a CDC-Anna 13a 3430a CDC-Chico 11a 2043c Myles 11a 2665b CDC-Nika 11a 3209a P value 0.5 0.0002 Standard error 1.49 153 Weeks after emergence-whole plant N N derived from Year Location Genotypes the atmosphere [N.sub.2] fixed % kg [ha.sup.-1] 2002 Saskatoon Amit 13c 11b CDC-Anna 26a 12ab CDC-Chico 16bc 13ab Myles 26a 15a CDC-Nika 22ab 14ab P value 0.007 0.1 Standard error 2.41 1.28 2003 Goodale Amit 29a 23bc CDC-Anna 25ab 28ab CDC-Chico 13c 12c Myles 28a 36a CDC-Nika 17bc 42a P value 0.01 0.007 Standard error 3.41 4.87 2003 SPG Amit 29c 33ab CDC-Anna 48a 37ab CDC-Chico 25c 21b Myles 31bc 48a CDC-Nika 46ab 46a P value 0.02 0.08 Standard error 5.3 6.6 ([dagger]) Means within a column followed by the same letter are not significantly different at P < 0.05.
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|Author:||Thavarajah, Dil; Ball, Rosalind A.; Schoenau, Jeff J.|
|Date:||Nov 1, 2005|
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