ACTA AGRONOMICA SINICA Volume 34, Issue 4, April 2008 Online English edition of the Chinese language journal

Cite this article as: Acta Agron Sin, 2008, 34(4): 609674. RESEARCH PAPER

Effects of High-Temperature Stress on Flowering and Grain-Setting Characteristics of Guodao 6

TAO Long-Xing, TAN Hui-Juan, WANG Xi, CAO Li-Yong, SONG Jian, and CHENG Shi-Hua*
National Key Laboratory of Rice Biology, China National Rice Research Institute, Hangzhou 310006, China

Abstract: Eight leading rice (Oryza sativa L.) hybrids were used to study the physiological and morphological adaptability to 4042C air temperature stress. Plants were treated for 6 h each day from initial heading to the 15th day. Guodao 6 and Xieyou 46, which had significant differences in heat injury index under high-temperature stress, were selected for investigating flowering and grain-setting characteristics response to high-temperature stress. Guodao 6 showed a relative stability in grain setting probably because of its adaptability to heat avoidance by changing characteristics of flowing. It had a shortened flowering period and weakened apical grain superiority under high-temperature stress. Within a day, although the peak flowering hour (11:00) was the same as control (normal temperature), the glume-opening rate decreased from 40.0% (control) to 23.6% (high-temperature treatment). In addition, the daily number of spikelet flowering increased compared with the control, and the flowering clock was delayed for 2 h during initial heading period. High temperature reduced the grain-setting rate of Guodao 6 by increasing the sterile spikelets. Keywords: high-temperature stress; heat tolerance; hybrid rice; flowering clock; glumes opening rate

The average daily temperature in the year 2030 is predicted to increase by 1.52.6C in China because of the aggravating greenhouse effect. In this condition, the fervent summer disaster with the maximum temperature over 40C and average daily temperature over 30C through half a month might occur frequently, which happened in the MiddleLower Reaches of Yangtze River in 2003 [18]. It is a challenge to rice cultivation associated with drought resistance and heat-injury resistance. Recently, some heat-tolerant rice (Oryza sativa L.) hybrids have been screened in Zhejiang Province, which was supported by a provincial project, including Guodao 6 and Zhong 9 you 8006. These hybrids have stable high grain-setting rate and spikelet fertility under high-temperature stress [9]. The objective of this study is to understand the physiological and ecological adaptabilities of rice cultivars under high-temperature stress (3035C [1014]), accordingly, to provide basic physiological parameters for heat-tolerance breeding in rice.

Xieyou 46, Liangyoupeijiu, Xieyou 9308, Zhong 9 you 8006, II you 7954, Zhongzheyou 1, and Shanyou 63, were studied at the experimental area of China National Rice Research Institute, Hangzhou, China. The seeds of 8 hybrids were directly planted in the fields on 25 May with the quantity of 10.512 kg ha1 based on the 1000-grain weight and germination rate. The seedling emergency and density were almost the same for each combination. Seedlings were then transplanted into pots for temperature treatments. In 2006, only 2 combinations, Guodao 6 and Xieyou 46, were used in a pot experiment. The pot (30 cm in diameter and 30 cm in height) was filled with 25 kg clayed soil from paddy field mixed with 0.025 kg commercial cake-fertilizer. The seeds were sown on 25 May, and only 2 healthy seedlings per pot remained for treatments. 1.2 High-temperature treatment

1
1.1

Materials and methods

Materials In the year 2005, 8 rice combinations, i.e. Guodao 6,

With reference to the methods of previous studies [1417], a modified heating and temperature control system was used in this study. The greenhouse was equipped with a set of far-IR Heating lamps (1000 W each lamp) and automatic temperature control system to control the temperature at 40 to 42C from 9:00 to15:00 every day. The high-temperature

Received: 1 August 2007; Accepted: 21 Nevember 2007. * Corresponding author. E-mail: shcheng@mail.hz.zj.cn Copyright 2008, Crop Science Society of China and Institute of Crop Sciences, Chinese Academy of Agricultural Sciences. Published by Elsevier BV. All rights reserved. Chinese edition available online at http://www.chinacrops.org/zwxb/

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treatment was started from the initial heading day and lasted for 15 d with 6 h (9:00 to 15:00) per day. Plants under the natural temperature were considered as control. 1.3 Investigation of grain-setting characteristics and heat injury index The 8 combinations were used to test the heat injury index in 2005. Plant samples were harvested at maturity with 4 replicates. Yield components, such as grain numbers per panicle, grain-setting rate, partially filled and infertile grains per panicle, 1000-grain weight, were investigated. The heat-injury index was defined as proportion of the difference of grain-setting rate between control and treatment to the grain-setting rate of control [9, 1619]. 1.4 Observation of flowering period and flowering clock

with the control, the grain-setting rate of heat treatment reduced, whereas the 1000-grain-weight increased. But the changing degrees were different among combinations. Guodao 6 showed the smallest heat injury among the 8 combinations. Under natural temperature (control), the spikelet fertility of Guodao 6 (spikelet sterility rate of 6.9%) was higher than that of Xieyou 46 (spikelet sterility rate of 12.2%). Under heat stress, Guodao 6 showed advantages in spikelet sterility rate and grain-setting rate than Xieyou 46. The heat injury index of Guodao 6 was significantly (P < 0.05) lower than that of Xieyou 46. These results indicate that Guodao 6 has better heat tolerance than Xieyou 46. The hightemperature treatment did not cause spikelet degeneration; therefore, total grain numbers per panicle change little in the 2 combinations (Table 1). 2.2 Effect of high-temperature stress on flowering period

Duration from the first spikelet flowering to the last spikelet flowering in a single panicle was defined as flowering period. Only Guodao 6 and Xieyou 46 were used to investigate their flowering period and flowering clock in 2006. Pots with growth uniformity of rice plant were selected and divided randomly into 2 groups: treatment and control. The treatments were given at meiosis stage of pollen mother cell. At the initial heading day, 10 uniform panicles were labeled in the morning, and the newly flowering spikelets were recorded each day for counting the anthesis period. These samples were harvested at maturity, and the yield component traits were recorded on a single panicle base. In this experiment, the average 1000-grain weights of Guodao 6 and Xieyou 46 were 31.4 and 25.2 g under natural temperature, respectively. Eighty percent and 10% of the values were used as criterions to classify fully filled (over 80%), partially filled (1080%), and infertile (below 10%) grains. In this experiment, the weights of partially filled grain (embryo formed but grain filled incompletely) were 3.224.9 mg for Guodao 6 and 2.518.9 mg for Xiyou 46. The flowering clock is the duration (hours) from the first to the last spikelet flowering in a single panicle on 1 day at full flowering stage. Usually, the flowering peak of singe panicle occurs at 34 d after initial heading, which is affected by panicle size and flowering period. In this study, 10 uniform panicles were selected for the observation of flowering clock on the 3rd day after initial heading. To facilitate the observation, the anthers of opened flowers (flowering at the 1st and 2nd days) were removed from the spikelets. The flowering clock was calculated by recording the number of blooming spikelets every hour till the end of anthesis [14].

Under heat stress, the glume opening rates of Guodao 6 and Xieyou 46 decreased by 4% and 6%, and the anthesis periods were shortened by 2 d (25%) and 1 d (17%), respectively. In both combinations, the blooming spikelet numbers of treatment were increased in the 1st day, and the anthesis peak appeared 1 day ahead of control (Table 2). 2.3 Effect of high-temperature stress on flowering clock

The flowering peak of Guodao 6 remained at 11:00 in high-temperature treatment, but the flowering clock was prolonged to 2 h compared with the control (Fig. 1). The number of flowering spikelets increased on the 1st day and the peak value decreased due to dispersed anthesis. The flowering rate was 40% under natural temperature at 11:00 but was only 23.6% under high-temperature stress. Similar phenomenon was observed in Xieyou 46, but the difference was not as significant as that of Guodao 6. 2.4 Effect of high-temperature stress on grain setting at different flowering period The apical grain superiority, i.e., superior spikelets of earlier anthesis and filling inhibited the inferior spikelets during grain-filling period, was weakened in both combinations under high-temperature stress, especially in Guodao 6 (Fig. 2). According to the average grain weight of Guodao 6, the spikelets flowering from the 1st to 5th day after initial heading were fertile under both control and high-temperature treatment (Fig. 2). Under natural temperature, the average grain weight of the 6th and 7th days were 23.0 and 17.5 mg, respectively, and the lowest value was 7.6 mg on the 8th day, which was only 23.3% of the highest value on the 2nd day (32.6 mg). However, when treated with high temperature, the lowest grain weight occurred at the 6th day with the average of 20.6 mg. This was accounting for 61.5% of the highest grain weight at the 1st day (32.5 mg).

2
2.1

Results
Effect of high-temperature stress on grain setting

The yield components of all combinations showed similar trends under high-temperature stress (Table 1). Compared

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Table 1

Yield components response to high-temperature stress from flowering to grain setting stage and heat injury index of 8 rice hybrids
183.1 4.9 a 182.2 11.2 a 164.2 7.2 a 160.7 8.3 a 185.3 4.1 a 187.6 1.2 a 179.9 4.5 a 201.3 4.4 a 143.4 7.6 a 142.5 12.0 a 190.0 8.3 a 189.6 12.9 a 180.0 9.2 a 178.2 11.3 a 137.8 7.6 a 136.7 12.0 a 52.1 4.6 b 84.4 1.3 a 33.2 3.2 b 79.2 4.4 a 37.4 3.2 b 81.4 3.6 a 42.6 4.6 b 83.3 4.1 a 48.0 4.3 b 81.5 4.3 a 34.2 0.9 b 78.5 4.3 a 47.6 0.4 b 87.5 0.6 a 45.1 0.3 b 80.3 0.3 a 36.7 1.32 b 8.7 0.42 a 41.6 2.01 b 8.6 0.52 a 48.6 3.00 b 11.3 0.35 a 41.8 0.69 b 8.6 0.35 a 41.7 1.20 b 11.3 0.72 a 44.4 2.90 b 11.2 0.51 a 39.2 0.61 b 8.6 0.72 a 43.9 0.31 b 11.8 0.54 a 11.2 0.14 b 6.9 0.05 a 25.3 0.21 b 12.2 0.61 a 14.4 0.19 b 7.3 0.41 a 11.6 0.75 b 8.2 0.91 a 10.3 0.46 b 7.2 0.35 a 19.4 0.51 b 10.3 0.07 a 13.2 0.71 b 8.9 0.82 a 11.0 0.51 b 7.9 0.70 a 32.0 0.31 a 31.4 0.71 a 26.3 0.27 a 25.2 0.26 a 25.9 0.17 a 25.6 0.07 a 25.9 0.17 a 25.6 0.07 a 26.2 0.09 a 25.7 0.54 a 27.4 0.28 a 26.5 0.61 a 27.0 0.32 a 25.6 0.71 a 27.0 0.28 a 25.7 0.38 a 0.445 c 0.423 d 0.564 a 0.411 d 0.487 c 0.546 b 0.581 a

Hybrid Guodao 6

Treatment Spikelets per panicle Grain-setting rate (%) Partially filled grains (%) Infertile grains (%) 1000-grain weight (g) Heat-injury index HT NT 0.382 d

Xieyou 46

HT NT

Liangyoupeijiu

HT NT

Xieyou 9308

HT NT

ZJY 8006

HT NT

II You 7954

HT NT

ZZY 1

HT NT

Shanyou 63

HT NT

In each hybrid, values followed by the same letter are not significantly different between the natural temperature (NT, control) and high-temperature treatment (HT) at the 0.05 probability level.

Table 2 Response of flowering period to high temperature in Guodao 6 and Xieyou 46

Hybrid Guodao 6 Treatment NT HT Xieyou 46 NT HT Spikelets per panicle 186.3 3.2 184.1 4.9 167.6 3.2 164.2 4.1 Glume dehiscence Number 184.4 175.2 161.1 148.1 Rate (%) 99.0 95.2 96.1 90.2 Flowering duration (d) 8 6 6 5 1st day 18.7 39.6 9.6 29.6 No. of glume dehiscence after n the day from heading 2nd day 40.3 41.3 39.8 46.4 3rd day 40.6 38.2 39.6 39.3 4th day 48.2 31.5 31.4 18.8 5th day 10.5 15.0 29.3 14.0 6th day 15.2 9.6 11.4 7th day 5.9 8th day 5.0

NT: natural temperature condition (control); HT: high temperature treatment. denotes data not available because of the termination of anthesis.

40

40

Guodao 6 Rate of glume dehiscence (%)

Xieyou 46

30

30

20

20

10

10

0 7:00 8:00 9:00 10:00 11:00 12:00 13:00 14:00 15:00 16:00

0 7:00 8:00 9:00 10:00 11:00 12:00 13:00 14:00 15:00 16:00

Flowering clock
Natural temperature

Flowering clock
High temperature

Fig. 1

Effect of high-temperature stress on spikelet flowering clock in Guodao 6 and Xieyou 46 Observed on the 3rd day from initial heading.

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Table 3 Effect of high-temperature stress on grain-setting rate (%) of spikelet in Guodao 6 and Xieyou 46
Hybrid Guodao 6 DAH 1 2 3 4 5 6 7 8 Xieyou 46 1 2 3 4 Fully filled grain Partially filled grain NT 100 98.7 100 90.0 81.2 70.4 23.0 0 100 90.6 78.4 72.3 60.2 0 86.5 80.2 65.2 40.3 9.2 HT 98.8 96.0 90.5 74.2 40.6 17.2 NT 0 0 0 0 8.8 24.1 61.3 88.6 0 0 10.0 10.3 16.0 77.6 0 3.8 20.3 30.4 57.6 HT 0 0 0 7.7 38.2 54.7 Unfilled grain NT 0 1.3 0 10.0 10.0 5.5 15.7 11.4 0 9.4 11.6 17.4 23.8 28.4 13.5 16.0 14.5 29.3 33.2 HT 2.0 4.0 9.5 18.1 21.2 28.1

Fig. 2 Effect of high-temperature stress on grain weight at different flowering time in Guodao 6 and Xieyou 46 NT: natural temperature; HT: high temperature.

5 6

DAH: days after heading; NT: natural temperature (control); HT: high temperature treatment.

In Xieyou 46, the average grain weight of spikelets flowering from the 1st to 4th day after initial heading exceeded the fertile criterion (20.0 mg) under both control and treatment (Fig. 2). The lowest grain weight was 5.0 mg on the 6th day and 12.5 mg on the 5th day in the control and high-temperature treatment, respectively. The lowest grain weights were 19.2% and 46.3% of the highest values in the control and high-temperature treatment, respectively. Table 3 showed that approximately 90% of spikelets flowering on the 1st to the 5th day in Guodao 6 developed either fully filled grains or partially filled grains under natural temperature. The infertile rates afterwards were ranged from 5.515.7%. But under high-temperature stress, the infertile rate on 4th day reached 18.1% and even rose up to over 20% on the 5th and 6th day. In Xieyou 46, the infertile rate was increased with the spikelet flowering time, and it was higher than that of Guodao 6.

Discussion

In the year 2003, grain-setting rates of some rice hybrids were below 30% in the middle and lower reaches of the Yangtze River area when encountered the strong heat stress in the summer. This again stimulated the researchers to focus on temperature resistant in rice. Most of the studies carried out in recent years focused on the creation and evaluation of the temperature-tolerant rice germplasm [1922]. In the year 20042006, the authors have detected heat tolerance in rice using interval-sowing method under natural and artificial high-temperature conditions. Guodao 6 was selected for its high heat tolerance and stable spikelet fertility probably associated with its restorer line [9]. The stable fertility of spikelet through the period from

anthesis to grain filling is suggested to be related to heat tolerance in rice [10, 13, 14, 19]. Two research directions on heat injury to spikelets were conducted in recent years. One was based on pollen viability indices, such as pollen size and pollen and stigma physiological activity related to grainsetting. The results showed that there was no significant correlation between I-KI or TTC dyeability rate and seed setting under high-temperature stresses at different stages. However, the pollen number and rate of pollen germination on stigma was significantly correlated with seed setting [17, 22, 23]. The other direction focused on analysis of dehiscence time and degree of anther with spikelet infertility. Significant correlation was found between the heat tolerance and the angle of glumes opening and number of pollen on stigma [18]. The authors suggested that daily maximum temperature had stronger impact on spikelet fertility than daily mean temperature [9]. In this study, compared with Xieyou 46, Guodao 6 showed a relative stability in grain setting under high- temperature stress, which may be associated with its adaptability to heat avoidance. Guodao 6 had approximately 200 spikelets per panicle, and normally, it took 8 d for complete anthesis. But it was shortened by 2 d under high- temperature stress. In Xieyou 46, only 1 day was reduced under the same stress (Table 2). The shortened period of anthesis leads to weaker apical grain superiority [2426] which facilitates the filling for the inferior spikelets. The weight ratio of early flowered superior grains to later flowered inferior grain was 100:23 in Guodao 6 under natural temperature, but it was 100:61 under heat stress. Similarly, the ratio increased from 100:19 to 100:46 in Xieyou 46. In addition to shortened period of anthesis, the flowering clock was also in response to the heat stress. Although the

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flowering peak was not advanced, the number of early flowered spikelets was increased at the 3rd day after initial flowering, especially the number of spikelets flowered at 7:008:00 in Guodao 6 (Fig. 1). The anthesis rates at peak hour (11:00) of Guodao 6 were 65% for natural temperature and 34% for high-temperature stress, indicating that high temperature prolonged flowering clock. A similar result was observed in Xieyou 46 but with less obvious differences between the control and heat stress treatment. Hu et al. [1] reported that low temperature prolonged flowering clock, which is in contrast to the result obtained in this study. The phenomenon requires further studies.

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Conclusions

Guodao 6 shows a relative stability in grain setting when treated with high temperature. The mechanism is probably based on the heat avoidance by shortening flowering period and dispersed flowering clock.

Acknowledgments
This study was supported by the Agricultural Structure Reform project of Ministry of Agriculture (06-03-01B), the Key R&D Project of Zhejiang Province (2005C22019 and 2008C22073), the National Agricultural Integrate Project (2006-27-2), and the Special Research Foundation for Commonwealth Institutes.

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