Source-sink balance and manipulating sink-source relations of wheat indicate that yield potential of wheat is sink-limited in high rainfall zones

Transcription

1 Source-sink balance and manipulating sink-source relations of wheat indicate that yield potential of wheat is sink-limited in high rainfall zones Heping Zhang 1,*, Neil C Turner 2, Michael L Poole 1 1 CSIRO Plant Industry Private Bag, PO Wembley, Australia, WA Centre for Legumes in Mediterranean Agriculture, Mail Bag M080, University of Western Australia, 3 Stirling Highway, Crawley, WA 6009, Australia * Corresponding author. Tel ; fax: address: heping.zhang@csiro.au 1

2 2 30 Abstract Grain yield depends on the number of grains per unit area (sink) and the availability of assimilates (source) to fill these grains. The aim of the current work was to determine whether wheat yield in the high-rainfall zone of south-western Australia is limited in current cultivar by the size of the sink or by the assimilates available for grain filling. Three wheat cultivars (Calingiri, Chara and Wyalkatchem) and two breeding lines (HRZ216 and HRZ3) were grown in four replicates in the field from 0 to 07. Dry matter and water soluble carbohydrates (WSC) at anthesis and maturity were measured and used to determine the source and sink balance of the crop. In 07, three further treatments were applied to manipulate the sink-source relationships: (i) Spikelets were removed on main stems to increase the source:sink ratio; (ii) The incoming solar radiation was reduced by 40% by shading after anthesis to reduce the availability of assimilates to grains; (iii) Supplemental irrigation was used to maintain the capacity for photosynthesis by an improved water supply during grain filling. The source-sink balance of the crops showed that the potential source was 2% greater than the actual grain yield in average and above-average seasons (0 and 07), suggesting that sink size, represented by the number of grain per unit area, was a limiting factor to yield potential. However, the source may have become a limiting factor in a drought season (06). The grain yield increased with increased number of grains/m 2 and kernel weight remained relatively stable even when grain number increased from 7000 to per m 2. The removal of half of the spikelets on the main stem did not increase kernel mass of the remaining grains and an additional 33 mm of irrigation water did not increase grain yield, but significantly (P < 0.0) increased WSC left in stems and leaf sheaths at maturity. Shading after anthesis did not significantly reduce grain yield of the current cultivars Calingiri and Wyalkatchem, but it reduced grain yield by 23-2% (P < 0.0) in Chara and HRZ3. All shaded plants extracted more WSC from the stem and leaf sheaths to partially compensate for the loss of assimilate production. The source-sink balance over three seasons and three independent experiments in 07 suggested that the yield of the current wheat cultivars is more sink- than source-limited and that breeding wheat with a larger sink size than in the current cultivars may lift the yield potential of wheat in the high-rainfall zone of south-western Australia. However, it is possible that once the sink size is enlarged, the yield potential of wheat could become source-limited. 2

3 Introduction Improvements in the yield of wheat are largely the result of increases in the number of grains per unit area (Fischer 07; Perry and D'Antuono 1989; Sayre et al. 1997) and partitioning of biomass to grains (higher harvest index) as a result of the introduction of dwarfing genes (Austin et al. 1980). Past experience in breeding and a lack of response of individual grain mass to the manipulation of sinks and sources have led crop physiologists to suggest that the grain yield of wheat is sink-limited and that increasing the sink capacity would increase the yield potential (Fischer 07; Miralles and Slafer 07; Reynolds et al. 07). The final grain yield can be considered as the balance between the supply of carbohydrate (source) and the capacity of the grains to accumulate available carbohydrates (sink). Carbohydrates for grain filling are supplied concurrently from photosynthetic activities and from reserves in the stem and leaf sheath that develop around anthesis. Sink capacity is a function of the number of grains per unit area and their potential size. Many source-sink manipulation experiments induced by degraining found either no increase in the weight of the remaining grains, or rarely the same magnitude as the increase in the source:sink ratio (Calderini and Reynolds 00; Calderini et al. 06; Ma et al. 199; Slafer and Savin 1994). Others, using defoliation (Kruk et al. 1997) and shading (Beed et al. 07), found that yields of wheat are co-limited both by the source and by the sink. Borras et al. (04) quantitatively determined the magnitude of changes in grain dry weight in response to manipulation of assimilate availability during grain filling and concluded that the yield of wheat is limited mainly by the size of the sink during grain filling. However, 2 many of these studies focused on the effects of the treatments on individual grain 3

4 weight, but not on final grain yield (Calderini and Reynolds 00; Calderini et al. 06; Ma et al. 199; Slafer and Savin 1994). The responses of individual grains in selected ears to the availability of assimilates may differ from the responses of grain yield per unit area at the field level to the same manipulation, and the field-level responses to sink and source manipulation need to be investigated. Furthermore, in the studies cited, no measurements were made to determine whether there were any water soluble carbohydrates (WSC) remaining in the plant at maturity, thereby indicating that there was an excessive source of assimilates in the plant during grain filling. The confirmation of an excess source at maturity would reinforce the hypothesis of sink limitation in wheat and provide evidence for increasing the sink capacity in breeding for increased yield potential. In southern Australia, wheat is traditionally grown in the low- and medium-rainfall areas with annual rainfall of less than 40 mm. Wheat yields of semi-dwarf variety in these areas are unlikely to be limited by sink size because terminal water stress during grain filling frequently forces rapid leaf senescence and reduces the assimilates available for grain filling. Wheat breeding targeting the low- and medium-rainfall zones of the cropping region has been focused on improving yield through early flowering (Regan et al. 1997; Siddique et al. 1989), high number of grains and associated high harvest index, and kernel weight for quality characteristics (Perry and D'Antuono 1989). As a result of low and erratically rainfall during grain filling period, the assimilates for filling grains are limited and therefore a large sink size may not necessarily be a preferred trait for this environment. There is a possibility that wheat breeding targeting the low- and medium-rainfall zones of the cropping region might 4

5 have unconsciously bred for limited sink capacity in order to avoid producing a large number of grains per unit area that cannot be filled due to a source limitation. The increasing profitability of annual cropping has brought a gradual expansion of wheat and other cereal crops into the more favourable high-rainfall zone of southern Australia (Zhang et al. 06). This region will become more and more important in grains production as climate change makes grain production in the low rainfall zone become marginable. In the high-rainfall zone, the yields of wheat that are currently achieved are only 0% of the potential yields (Anderson ; Zhang et al. 06) and 1-2 t/ha lower than the simulated yield predicted by the APSIM wheat model under well managed field conditions (Zhang et al. 07). Apart from constraints such as waterlogging during the winter months, low nutrient inputs and poor weed management, a lack of cultivars adapted to the longer growing season and higher and more reliable rainfall is an important constraint in the high-rainfall zone (Zhang et al. 04; Zhang et al. 06). Sink limitation has been reported to be a limiting factor to yield potential of wheat and barley in the environments with the absence of serious post-anthesis stress (Beed et al. 07; Bingham et al. 07; Borras et al. 04). The wheat cultivars currently grown in the high-rainfall regions are those bred for the lowand medium-rainfall zones. These varieties developed for and well-adapted to the low and medium rainfall region may be able to produce a large enough sink size to accommodate the available assimilates produced in the water-limited environment. However, when these wheat varieties are grown under the more favourable rainfall zones of southern Australia, their sink size may be limiting the potential yield in the region. 2

6 The study reported here was designed to: i) determine the source:sink relationships in the current cultivars; and ii) examine the impact of manipulating the source:sink ratio for assimilates on the yield of wheat during grain filling in the high-rainfall zone of south-western Australia. The aim of the study was to determine whether the yield of wheat in this zone is limited by the sink size of the current varieties, or by the potential assimilates available for grain filling, or co-limited by sink and source, and thereby the implications for the breeding of new wheat cultivars for the high-rainfall regions of southern Australia. Methods and materials Site, experiment and design The experiments were conducted in 0 to 07 on a farm km south of Kojonup in Western Australia (33 o S, 116 o 4 E). The site was equipped with an automatic weather station to monitor the minimum and maximum temperatures, solar radiation. Daily rainfall was recorded using a tipping bucket rain-gauge connected to the weather station. The soil was a sandy duplex soil with 0.4 m of gritty loamy sand overlaying clay. The source and sink balance experiment Three commercial spring wheat (Triticum aestivum L.) cultivars Carlingiri, Wyalkatchem, and Chara and one breeding line (HRZ216 in 0 and 06, and HRZ3 in 07) were sown at a seeding rate of 0 kg/ha on 30 May 0, 23 June 06, and 23 May 07. The established plant density was around 1-140, , plants/m 2 in 0, 06, and 07, respectively. Wyalkatchem and Calingiri 2 were selected to represent the commercial cultivars grown in the region because 4% 6

7 of the area in Western Australia was sown to these two cultivars from 04 to 07 (Zaicou et al. 08). Chara and the two breeding lines, HRZ216 and HRZ3, were selected because they had the capacity to produce more grains per unit area due to their large head size compared to the two commercial cultivars. HRZ3 was added in the experiment in 07 because of its longer growing season and high yield potential than HRZ216. A randomized complete block design was used with four replicates. Each plot was 30 m by 2.88 m (16 rows, 0.18 m apart). In all three years, the wheat crop was supplied with 1 kg N/ha split as follow: kg/ha at sowing, 0 kg/ha at tillering, and 0 kg/ha at stem elongation. Seventeen kg P/ha was drilled into the soil at sowing as mono-ammonium phosphate and 0 kg K/ha in form of muriate of potash was top-dressed along with N at tillering. The crop was sampled using a quadrat of 1.08 m 2 to determine dry weight and water soluble carbohydrates in stems and leaf sheath at anthesis and physiological maturity each year (see crop measurement for details). The difference of dry weight between anthesis and physiological maturity plus WSC stored in stem and leaf sheath at anthesis was taken as the potential source available for grain filling. The final grain yield was assumed to be the sink size. This is similar to the method used by Bingham et al. (07) in analysing the source-sink balance. It is acknowledged that postanthesis dry matter accumulation can be positively associated with the number of grains and can be inhibited by a feedback from a limited sink capacity (Bingham et al. 07; Reynolds et al. 07; Slafer et al. 1996). However, this feedback inhibition from a limited sink size does not affect our analysis because the potential source would have be even greater than the observed post-anthesis dry matter if the sink size 2 was larger. Therefore, comparison of the sizes of source and sink and the residual 7

8 amount of WSC in stems and leaf sheath at maturity were used to determine the limitation of source and sink. Spikelet removal experiment In 06 and 07, immediately after each cultivar reached 0% anthesis, 6 pairs of main stem ears (a total of 12 ears) were selected and tagged in each plot of Calingiri, Wyalkatchem, Chara, and HRZ216 (06 only) and HRZ3 (07only). The paired ears had the same number of spikelets. All spikelets from one side of the six tagged ears were removed by hand and the other six ears left intact as a control. The removal of half of the spikelets had the potential to double the assimilate supply to the remaining spikelets in those spikes. At maturity, the paired ears were harvested separately and hand threshed. The number of spikelets and grains per ear were counted. Kernel weight was measured by dividing the grain weight by the number of kernels per ear. Shading experiment In 07, a shading experiment investigated the effect of reducing incoming radiation during the post-anthesis period on grain yield and yield components and evaluated the effect of reducing assimilates supply from photosynthesis on grain size and the amount of WSC remaining at maturity. The shading was imposed on the four genotypes (Wyalkatchem, Calingiri, Chara and HRZ3) using horticultural shade cloth immediately after the crop reached 0% anthesis. An area of 1. m by 1.8 m was shaded in each plot. The shade cloth was supported by four star picks hammered into ground with a height of 1.60 m above-ground. The cloth covered the top of the 2 designated area and extended downwards 1.2 m from the top from all directions 8

9 except the southern direction. The photosynthetically-active radiation (PAR) measured from :00 h to 16:00 h on three days during the grain filling period showed a 40 2% reduction of incoming PAR. Supplemental irrigation experiment In 07, an irrigation experiment investigated the effect on grain yield of providing additional water to the crop during grain filling and the amount of WSC remaining at maturity. The supplemental irrigation was applied on micro-plots in each plot of four cultivars (Wyalkatchem, Calingiri, Chara and HRZ3) days after 0% anthesis. The micro-plots consisted of an area of 1.80 m by 1.2 m. The irrigation was applied by hand using watering cans with a known volume. The total water applied was estimated using a soil water balance model to ensure that the ratio of pre- to postanthesis water use was at 2:1 (Passioura 1983; Zhang et al. 0). The water applied was 33 mm. Crop measurements Crop growth stages were scored according to Zadoks et al. (1974). Crop samples were collected using a quadrat of 1.08 m 2 (1-m length of row from 6 adjacent rows). At anthesis, one sample was taken from each plot. At maturity, samples were taken for control, shaded, and irrigated treatments from each plot. Ten plants were randomly sub-sampled and partitioned into leaves, stems with leaf sheath, and ears. All samples were dried to constant weight in a forced-draught oven at 60 o C and weighed. At anthesis, leaf area was measured using a Licor area meter (LI-3000A, Licor, Lincoln, NE, USA). 2 9

10 Grain yield was determined from the 1.08 m 2 quadrat cuts from each plot. All ears from the area sampled were counted to estimate ears/m 2 for each plot. Ten plants were then randomly chosen from each sample and threshed by hand and the number of grains counted to determine grains/ear. Kernel weight was measured using one thousand kernels. The number of grains/m 2 was calculated by dividing the grain yield by kernel weight. Water soluble carbohydrates In all three years, five plants from the quadrat samples at anthesis and physiological maturity were randomly selected and separated into leaf blades, stems with leaf sheath, and spikes. All samples were dried at 60 o C for 48 h and weighed. The stem samples were ground and passed through a 1 mm sieve and used to determine the concentration and amount of WSC in the stems at anthesis and maturity. WSC were extracted from 0.1 g of ground stem material by extracting once with 8 ml of 80% ethanol at 80 o C followed by 2 extractions with 8 ml distilled water at 60 o C. Each extraction was 3600 s. The extraction was centrifuged at room temperature for 600 s at 3400 rpm and the extracts were combined. Total carbohydrates in the samples were analysed by the anthrone method (Yemm and Willis 194). Results Seasonal conditions In 0, the annual rainfall of 76 mm at the site was significantly higher than the long-term average at Kojonup (34 mm), with growing-season rainfall of 26 mm. It represented a wet year in the region (top percentile of the seasons). Year 06 was 2 a drought year with 360 mm annual rainfall of which 296 mm fell during the growing

11 season. The growing season rainfall in 06 fell in the percentile of 2% (one in every four year). The drought early in the season delayed sowing until late June. Year 07 represented an average year with annual rainfall of 40 mm and growing-season rainfall of 396 mm. Three frost events occurred during and after anthesis in 0, significantly reducing the number of grains per ear of the genotypes. There was no frost event around anthesis in 06 and 07. Sources and sinks of wheat The wheat crops achieved more than g/m 2 of dry matter at anthesis in 0 and 07 and set up the crops for high yields of g/ m 2 (4-6 t/ha) (Table 1). In 06, lack of water combined with late sowing halved the dry matter production at anthesis compared with 0 and 07. The difference in dry matter production at anthesis also resulted in the amount of WSC and post-anthesis dry matter accumulation being halved in 06 compared to 0 and 07. We assume the g/m 2 of WSC remained in the stems and leaf sheaths at maturity represents full remobilization of WSC although the absolute amount of WSC can vary with agronomic management (van Herwaarden et al. 1998) and the post-anthesis growing conditions (Ruuska et al. 06). This value was based on the estimation of g/m 2 of stems and leaf sheaths weight at maturity for a yield of 6 t/ha (Zhang et al. 07) and their WSC concentration of 4-% (Ruuska et al. 06). This value is also similar to the reported values of 2 for winter wheat (Foulkes et al. 07) and 27 g/m 2 for barley (Bingham et al. 07) in the high yielding (8- t/ha) UK environment, and g/m 2 for wheat observed in Australian (van Herwaarden et al ). The sum of the post-anthesis dry matter accumulation and the WSC stored in 11

12 stems and leaf sheath at anthesis above the g/m 2 of WSC threshold was defined as the potential source for filling grains and achieving the potential yield. The amount of WSC remained in stem and leaf sheath at maturity varied between the seasons and between cultivars within the same seasons. In 06, all the wheat genotypes extracted all the available WSC and remobilized all the potential sources for grain filling (Table 1). In 07, Wyalkatchem left significantly more WSC at maturity than other three genotypes, yielding about 30% less than the potential source (Table 1). In 0, the frost around and after anthesis reduced the number of grains (sink size). The wheat genotypes with the reduced number of grains in this season can be seen as ones with a limited sink size. More than a half of stored WSC at anthesis remained in stems and leaf sheaths of Wyalkatchem and Calingiri at maturity. Grain yield was positively related to the number of grains/m 2 (r 2 =0.81) (Figure 1a). The WSC remaining in the stems at maturity was negatively related to the number of grains/m 2 at maturity in 0 and 07 (r 2 =0.9), but not in 06 when the source become limited due to late planting and drought (Figure 1b). The negative relationship indicates that the fewer grains/m 2 (small sink size) may have contributed to the lack of remobilization of the WSC. If the wheat had a sufficient number of grains and was able to remobilize the available WSC to the grains, the calculated potential yields in average and above average years (0 and 07) approached 0-70 g/m 2 (.-7. t/ha) and the potential harvest index was about 0.0 (Table 1). Relationship of grain yield and kernel weight to grain number In order to create a wide range of responses of grain yield to grain number and kernel weight to grain number, the data from additional experiments at Kojonup were used to 2 examine the effect of variation in grain number on grain yield and kernel weight. For 12

13 Wyalkatchem, the data in Table 1 along with the previous experiments for Wyalkatchem from 01 to 03 (Zhang et al. 07) were combined. For Calingiri, the data in Table 1 were combined with separate experiments conducted in 06 and 07 involving three seeding rates and four nitrogen levels (H. Zhang, 09, unpublished). Grain yield increased with increasing number of grains/m 2 in Wyalkatchem and Calingiri (r 2 = 0.94) with no significant difference in the slopes (Fig 2a), while kernel weight showed no apparent decrease with increasing grain number in Wyalkatchem (Fig. 2b) and a slight decrease in Calingiri (Fig. 2b) (P < 0.0). There was a doubling of yield for a doubling of grain number from about 300 to 600 g/m 2 in both cultivars, whereas the change of kernel weight was less than % when the grain number doubled. Responses to spikelet removal The removal of spikelets on one side of the spikes halved the number of spikelets and grains per spike in both years (Table 2). This potentially doubled the resources available to the rest of the grains in the spikes. However, this did not increase the kernel weight of the remaining grains in 07 (Table 2). It was surprised to note that kernel weight was reduced by spikelet removal except Wyalkatchem in 06. While the increase in kernel weight was between 8 and 13% in Chara and HRZ3, this was not statistically significant, and the increase of kernel weight in Calingiri and Wyalkatchem was < % compared with the controls in 07. The kernel weight of Calingiri and Wyalkatchem was significantly higher than that of Chara and HRZ3. Responses to additional water supply during grain filling 13

14 All genotypes reached 0% anthesis within about 12 days (Table 3) each other. Wyalkatchem flowered first, followed by Calingiri and then Chara. The main stem of HRZ3 flowered as early as Wyalkatchem, but the spikes from tillers reached 0% flowering as late as Chara. The leaf area index (LAI) was about 1. for all genotypes except HRZ3 which had a LAI of 2.2. The soil moisture content in the top 40 cm soil was around 16% for Calingiri, Chara and HRZ3, and % for Wyalkatchem just before the irrigation treatments were applied. Wyalkatchem and Calingiri received 6 mm more post-anthesis rainfall than the other two genotypes because of earlier anthesis. For the unirrigated controls, the number of grains/m 2 varied significantly between the cultivars. Chara and HRZ3 had a larger sink size at 4000 more grains/m 2 than Calingiri and Wyalkatchem (Tables 1 and 4). The additional 33 mm of water supplied by irrigation did not affect the total dry matter, ears/m 2 and kernel weight, but increased the number of grains/ear from 39 to 42 (P < 0.01) (Table 4). This increase did not result in a significant or consistent increase in grains/m 2 or grain yield. The additional water supply significantly (P < 0.0) increased the WSC content of the stems and leaf sheaths on average from.3% to 17.% and the absolute amount of WSC from 1 to 90 g/m 2 in stems at maturity (Table 4). The stored WSC in stems ranged from 190 to 243 g/m 2 at anthesis and from 22 to 6 g/m 2 at maturity. The difference of WSC between anthesis and maturity was 13 to 213 g/m 2. The WSC concentration in stems at maturity was in a range of -9% in Calingiri, Chara and HRZ3 in the unirrigated plots, indicating that the majority of stored WSC at anthesis was transferred to grains in those cultivars. However, in Wyalkatchem, significantly more (P < 0.0) stored WSC was left in the stems and 2 leaf sheaths at maturity than in the rest of cultivars (Table 4). In the unirrigated plots, 14

15 the stored WSC contributed 2% to grain yield in Calingiri and Wyalkatchem, and 36-40% in HRZ3 and Chara, but contributed about % and 30% when supplied with additional water, respectively. Responses to shading during the post-anthesis period Shading after anthesis significantly reduced the total dry matter at maturity from 1340 to 1190 g/m 2 (13.4 to 11.9 t/ha) (P < 0.01), reduced grains/m 2 from 147 to 130 (P < 0.0) and kernel weight from 38.6 to 36.6 mg (P < 0.0), and caused grain yield to decrease from 7 to 471 g/m 2 (.7 to 4.71 t/ha) (P < 0.01) (Table 4). However, the response in dry matter and grain yield to shading differed among the genotypes. Dry matter, ears/m 2, grains/m 2 and grain yield of Calingiri and Wyalkatchem were not significantly affected by shading, whereas shading significantly reduced dry matter, grains/m 2 and grain yield of Chara and HRZ3, primarily because of slight but not significant decrease in the number of ears/m 2. The number of grains/ear in all genotypes was not affected by shading. The shaded wheat did not extract significantly more WSC from the stems and leaf sheaths compared to the unshaded wheat. Discussion Source-sink balance The comparison of the potential assimilates (source) for grain filling with the actual yield of wheat crops suggests that the source was significantly greater than sink in the seasons with average rainfall (07) and above-average rainfall (0) and that the wheat crops with limited sink size left a considerable amount of WSC in stem at maturity. Our results showed the potential assimilates available for grain filling would 2 have increased yields by 2% over the actual yields and the harvest index would have

16 been 0.0 if the current wheat cultivars were not constrained by the limited number of grains (sink capacity). (Table 1). Limited sink size has been reported to inhibit current photosynthesis of crops as a result of feedback from abundant assimilates for grain filling (Bingham et al. 07; Reynolds et al. 07; Slafer et al. 1996). Therefore, the potential source may have been even greater than we observed. If the sink size of the current cultivars can be increased, it is possible that similar harvest indexes could be achieved in the high-rainfall zone of south-western Australia to those in other high yielding environments (Foulkes et al. 02; Shearman et al. 0). The source sink balance indicated that extras sources remained in plant at maturity were stored either in stems and leaf sheaths as a form of WSC (Table 1) and/or in chaff (a net increase in chaff weight in 0) (data not showed). However, seasonal growing conditions can influence the limitation of sources and sinks. The full remobilization of available WSC in 06 suggests that the available assimilates were limited in this drought season. Fischer and HilleRisLambers (1978) have reported that seasonal conditions such as temperature, solar radiation, drought can impact on variation in source-sink limitations. As the percentile of rainfall in 06 occurs in only one in four years, we conclude that in most seasons the yield potential of wheat is limited by sink size rather than the potential source of the current cultivars in the high-rainfall zone of southwestern Australia. Responses of grain yield to grain number and kernel weight The positive linear relationship between grain yield and grain number and lack of decrease in kernel weight with increasing grain number in Wyalkatchem and Calingiri (Figure 2) reflect that the source from the current photosynthesis and stored WSC in 2 stem and leaf sheath was relatively abundant and that the inter-grain competition was 16

17 small in these two cultivars. This conclusion is also supported by our observation of no significant reduction in grain yield in response to shading in these two cultivars (Table 4). The lack of a significant decrease in kernel weight in these two cultivars (Figure 2) is in contrast with the observation made by Fischer et al. (1977) who found that kernel weight fell linearly with increase in grain number over a wide range of grain numbers for high-yielding wheat. However Chara and HRZ3, the genotypes of wheat with a greater number of grains/m 2 than Wyalkatchem and Calingiri showed a similar response to that reported by Fischer et al. (1977) and exhibited some source limitation as kernel weight decreased with increased grains/m 2 (Tables 1 and 3). The full extraction of WSC in Chara and HRZ3 also indicated that assimilates may have been limited in these two genotypes with high grain number. This suggests that the yield could have become source-limited when sink size was large. Although kernel weight fell linearly with increase in grain number, Fischer et al. (1977) found that grain yield increased with increased grain number and reached a maximum at grain numbers well above the levels of control crops. Considering the limited decreases of kernel weight in Wyalkatchem and Calingiri, we consider that increasing number of grains per unit area is an avenue to increase the yield potential of wheat in highrainfall environments even if kernel weight may be reduced in some seasons and some cultivars. Responses to alteration of sink and source Altering the source:sink ratio either up or down through providing extra water to crops and shading during grain filling provides further evidence for our hypothesis that the wheat yield of the current cultivars is more sink- than source-limited in the 2 high-rainfall zone of south-western Australia. The lacking of increases in kernel 17

18 weight to spikelet removal in 06 appeared to contradict to our conclusion of source limitation as a result of drought from the source-sink balance analysis in that year. This suggests that spikelet removal may be flawed in quantifying source-sink limitation. It is also possible that delaying of sowing from the drought in 06 reduced pre-anthesis growth and thus the sink size while there was adequate assimilates supply to fill this limited number of grains. Nevertheless, increasing the source-sink ratio by removing a half of spikelets of the spikes in 07 did not increase the kernel weight of the remaining grains (Table 2). This is consistent with previous work in wheat where individual kernel weight was rarely reported to be limited by the source during grain filling in other high-yielding environments (Calderini and Reynolds 00; Calderini et al. 06; Ma et al. 199; Slafer and Savin 1994). This is also consistent with our conclusion from the source-sink balance analysis in 07. The kernel weight of the main stems of Wyalkatchem achieved in 07 (Table 2) was % higher than the kernel weight achieved in well-fertilised and well-watered Wyalkatchem grown in the glasshouse (Robertson et al. 09). Considering that the kernel weight of spikes from the main stem was greater than that of spikes from tillers, the kernel weight of the cultivars in this study may have approached their geneticallycontrolled maximum kernel size (Fischer and HilleRisLambers 1978). This leads us to conclude that there is little likelihood of increasing grain yield by further increasing kernel weight (size) in the high-rainfall zone of south-western Australia. The lack of response of grain yield to the additional water supply and the increase in WSC in the stems and leaf sheaths at maturity under irrigation also imply that there was insufficient sink capacity in the current wheat cultivars. At maturity, the 2 concentration of WSC in Wyalkatchem and Calingiri under non-irrigated conditions 18

19 and in all genotypes under irrigation was much higher than the % threshold that represents full remobilisation of stored carbon (Ruuska et al. 06) (Table 2). In particular, Wyalkatchem left more g/m 2 of WSC in the stems and leaf sheaths at maturity when the number of grains was limited. This indicates that the current assimilates were sufficient to maintain the demands of grain growth and that yield of wheat was not limited by assimilate supply. On the other hand, the lack of a reduction of grain yield in Calingiri and Wyalkatchem by post-anthesis shading confirms that the assimilate availability in the shaded plants plus the WSC at anthesis was sufficient to meet the demand for grain growth. This provides further evidence for abundant assimilates for grain filling in the current cultivars. Shading reduced post-anthesis dry matter accumulation by 18-%, but the reduced assimilate supply from current photosynthesis was partially compensated by more extraction of WSC from stems (Table 4). This resulted in only a % yield reduction for Wyalkatchem and 9% for Calingiri. However, the impact of shading on Chara and HRZ3 was significantly higher than that of Wyalkatchem and Calingiri, significantly reducing post-anthesis dry matter accumulation, kernel weight and grain yield (Table 4), and indicating that assimilates may have become limiting when the grain number was increased. With shading, the calculated source limitation factor (the ratio of change in grain yield relative to change in total dry weight, (Gifford et al. 1973) was 0.2, 0., 0.4 and 0.7 for Wyalkatchem, Calingiri, HRZ3 and Chara, respectively. The smaller value for Wyalkatchem indicates that it is more sink-limited and the higher value for Chara suggests that it is more source-limited. Calingiri and HRZ3 could have been equally sink- and source-limited. The grain yield of wheat 2 has frequently been reported to be equally source- and sink-limited (Fischer 197; 19

20 Gifford et al. 1973; Kruk et al. 1997). The lack of decreases in kernel weight and grain yield of Wyalkatchem and Calingiri contrasted with significant reductions of kernel weight and grain yield in Chara and HRZ3. The difference in the responses to the manipulation of sink and source between the genotypes suggests that once the sink size is increased, yield could become source-limited in the high rainfall zone of south-western Australia. Breeding implications This study has clearly shown that the yield of wheat in the current cultivars is more likely sink-limited than source-limited in the high-rainfall zone of south-western Australia. Wyalkatchem and Calingiri, originally bred for the low- and mediumrainfall regions of the cropping zone, may have sink size limitations when grown in the high-rainfall zone. It is possible that in selecting for large seed size to avoid large screenings of pinched grain as a result of terminal water stress, breeders selecting cultivars for the low- and medium-rainfall zones may have inadvertently reduced the seed number. These cultivars bred for the lower rainfall areas appear to produce a maximum number of seeds of grains per m 2 (Figure 2). Furthermore, since wheat breeding in Western Australia has been heavily targeted towards the lowand medium-rainfall zones, parents with large sink size and shrivelled grain when exposed to terminal drought are likely to be considered inferior and discarded. We suggest that is the selection of genotypes for the medium- and low-rainfall areas has limited the potential yields of the current wheat cultivars when grown in the highrainfall zone of south-western Australia. This study has shown that the amount of assimilates of current cultivars was abundant and the level of competition between 2 grains for available assimilates was low when grown in the favourable climate of

21 south-western Australia. Therefore, we suggest that selection for increased grain number per unit area could lead to an increase in yield potential. Sink size may be increased by increasing the crop growth rate during spike growth period by increasing resource capture or radiation use efficiency (Fischer 07), and lengthening the duration of the spike growth phase through manipulation of the sensitivity to photoperiod (Miralles and Slafer 07). In the high-rainfall areas of south-western Australia, the wheat cultivars currently available cannot be sown in late April or early May in years when soil moisture is plentiful because of a high frost risk at anthesis in mid- and late-september. We previously found that the availability of water during the post-anthesis period in the high-rainfall zone of south-western Australia can allow wheat cultivars to have a flowering date days later than Wyalkatchem without prematurely exhausting soil water (Zhang et al. 0). The early sowing opportunities and a -day delay in flowering provide avenues for breeders to extend the length of spike growth period. The flowering dates of the current cultivars varied over a -to -day period depending on the time of sowing, but the actual length of period from terminal spikelet initiation to flowering was similar (around 6 days) among the cultivars (data not shown). This suggests that there is a lack of genetic variation in the length of the spike growth period among the spring wheat cultivars in this study. It would be valuable to examine the genetic variation in the length of spike growth period in wheat and the relationship between it and the sink capacity. The ideotype of wheat for the high-rainfall zone of southern Australia is one that initiates reproductive growth (double-ridge formation) as early as Wyalkatchem and flowers at a similar time to Calingiri so that the length of the spike growth period is increased. 21

22 The other way to increase sink capacity is to examine the genetic variation in grains per unit of spike weight (Fischer 07). Our study indicates that genetic variation in grains per unit of spike weight already exists among the current wheat cultivars, as Calingiri and Chara produced 0 grains per unit spike weight, whereas Wyalkatchem and HRZ3 produced only 70 grains per g spike dry weight. No relationship was found between the grains per unit spike weight and the number of grains, but as this study involved only four cultivars, it is clear that further studies are warranted to explore the genetic difference in grains per unit spike weight. High ear number has been reported to be the key to achieving high yield in the current wheat cultivars in the high rainfall zone of Western Australia and the current cultivars have the capability to reach the number of ears per unit area required for great than 6 t/ha (Zhang et al. 07; Zhang et al. ). This probably suggests that increasing the number of grains per ear may provide avenues to increase the sink size of the current wheat cultivars. In conclusion, the experiments conducted in this study on three commercial cultivars and a breeding line have shown that the yield potential of wheat in the high-rainfall region of south-western Australia is more likely to be limited by sink capacity rather than source availability. Therefore, increasing the sink capacity of wheat by increasing the number of grains per unit area should lead to higher yields and the realisation of potential yields in the region. Acknowledgements 22

23 We thank Tammi Short, Zhenhua Zhang and Justin Laycock, for data collection, Kelly Whisson for analysing the water soluble carbohydrates, and Vince Lambert at the Western Australia Department of Agriculture and Food at Katanning for agronomic management of the trials. The breeding lines HRZ216 and HRZ3 were provided by Dr Richard Richards of CSIRO Plant Industry. We acknowledge the Kojonup Crop Research Group for helpful discussions on the project and Peter and Anna Macleay for providing sites for the experiments. The project was supported by CSIRO and the Grains Research and Development Corporation. We thank Drs Jairo Palta and Steve Milroy for comments on an earlier draft of the paper. 23

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27 Table 1 Dry matter at anthesis (DW a ), post-anthesis dry matter accumulation (DW pa ), derived as increase in above-ground dry matter from anthesis (DC6) to physiological maturity, water soluble carbohydrates stored in stems and leaf sheaths at anthesis (WSC a ) and at physiological maturity (WSC m ), grain yield, yield components and harvest index (HI),and the estimated potential yield (Pot. Yield) and potential HI (Pot. HI) for three wheat cultivars and one breeding line in 0, 06, and 07. The Potential Yield = DW pa + WSC a -; Potential HI = Potential yield/dw. Cultivar DW a DW pa WSC a WSC m Grains/m 2 Kernel weight Yield HI Pot. Yield Pot. HI (g/m 2 ) (g/m 2 ) (g/m 2 ) ( 3 ) (mg) (g/m 2 ) (g/m 2 ) 0 Calingiri Chara Wyalkatchem HRZ l.s.d. (P = 0.0) n.s. n.s Calingiri Chara Wyalkatchem HRZ l.s.d. (P = 0.0) n.s. 3 n.s. n.s. n.s. 1.9 n.s Calingiri Chara Wyalkatchem HRZ l.s.d. (P = 0.0) n.s. 216 ns n.s

28 Table 2 Effect of spikelet removal on the number of spikelets, grains/ear and kernel weight in three wheat cultivars and one breeding line at Kojonup, Western Australia, in 06 and 07. Treatment/ Genotype Spikelets Grains/ear Kernel weight (mg) 06 Control Calingiri Chara HRZ Wyalkatchem Mean Spikelet removed Calingiri Chara HRZ Wyalkatchem Mean l.s.d. (P < 0.0) treatment l.s.d. (P < 0.0) cultivar n.s l.s.d. (P < 0.0) interaction Control Calingiri Chara HRZ Wyalkatchem Mean Spikelet removed Calingiri Chara HRZ Wyalkatchem Mean l.s.d. (P < 0.0) treatment n.s. l.s.d. (P < 0.0) cultivar l.s.d. (P < 0.0) interaction

29 Table 3 Date of 0% anthesis, above ground dry weight (DW a ), leaf area index (LAI), spike weight of three cultivars and one breeding line at anthesis, soil moisture content at anthesis before shading and irrigation treatments were applied, and post-anthesis rainfall at Kojonup, Western Australia, in 07. Genotype Anthesis DW a LAI Spike weight Soil moisture Rainfall after anthesis (g/m 2 ) (g/m 2 ) (cm 3 /cm 3, %) (mm) Calingiri 8// Chara 16// Wyalkatchem 6// HRZ3 18// l.s.d. (P = 0.0) - n.s

30 Table 4 Effect of additional water supply and shading during grain filling on total dry matter (DW), grain yield, yield components, and water soluble carbohydrates (WSC) in stems and leaf sheaths at anthesis and maturity in three wheat cultivars and one breeding line at Kojonup, Western Australia, in 07. Treatment/Genotype DW Ears Grains Grains/m 2 Kernel Yield WSC /m 2 /ear weight Anthesis Maturity Anthesis Maturity (g/m 2 ) ( 3 ) (mg) (g/m 2 ) (%) (g/m 2 ) Control Calingiri Chara HRZ Wyalkatchem Mean Irrigated Calingiri Chara HRZ Wyalkatchem Mean l.s.d.(p < 0.0) irrigation n.s. n.s n.s. n.s. n.s l.s.d.(p < 0.0) cultivar n.s l.s.d. (P < 0.0) interaction n.s. n.s. n.s. n.s. n.s. n.s. - n.s. - n.s. Shaded Calingiri Chara HRZ Wyalkatchem Mean l.s.d. (P < 0.0) shading n.s n.s n.s. - n.s. l.s.d. (P < 0.0) cultivar l.s.d. (P < 0.0) interaction 8 n.s. n.s. 2.2 n.s n.s. - n.s. 30