Effect of High Temperature during the Seed Development on Quality and Chemical Composition of Chili Pepper Seeds

Transcription

1 Jpn. J. Trop. Agr. 51(1) : 22-29, 2007 Effect of High Temperature during the Seed Development on Quality and Chemical Composition of Chili Pepper Seeds Piyanath PAGAMAS and Eiji NAWATA* Laboratory of Tropical Agriculture, Graduate School of Agriculture, Kyoto University Abstract An analysis was performed on chili pepper [Capsicum annuum (L.) cv. Huay Si Thon and Shishito] seed vigor and quality produced by plants grown under control and high temperature conditions (29 } 2/24 }2 and /27 } 2 Ž, mean day and night temperatures, respectively) after anthesis to elucidate the effect of high temperature during seed development on seed quality. High temperature significantly reduced fruit growth in both varieties. More than 20 % of the seeds produced under the high temperature regime were flat with a dark brown color and did not germinate. High temperature inhibited seed fresh and dry weight increase and the seed size was slightly reduced. Standard germination of Huay Si Thon and Shishito seeds occurring under the high temperature conditions was lower than that of the control seeds by 28 and 25 %, respectively. Similarly, seed vigor was reduced, as evidenced by the reduction in the accelerated aging germination rate and the higher value of the germination index. Protein content was maximal during the early stages of seed development and decreased afterwards continuously during the seed maturation stage, while no conspicuous reduction of the protein content by heat was observed. High temperature reduced the carbohydrate content of seeds by 40 % in Huay Si Ton and 50 % in Shishito, respectively. High temperature reduced the lipid content to less than half. These results suggested that high temperature during seed development reduced chili pepper seed germination and vigor, presumably due to the limitation in the accumulation of storage products, especially carbohydrates, and lipids. Key Words: Accelerated aging, Carbohydrate, Germination, Heat stress, Lipid Introduction Chili peppers grow best and are likely to reach maximum yields in the dry season at temperatures ranging from 21 to 33 Ž. However, because of environmental fluctuations, temperatures are often higher than the optimum ones (Khah and Passam, 1992), thus increasing the probability of the plant being exposed to extended periods of supra-optimum temperatures. In general, high temperature may lead to significant losses in crop productivity in many species due to limited vegetative and reproductive growth and seed yield (Cross et al., 2003; Erickson and Markhart, 2002; Monjardino et al., 2005; Spears et al., 1997). In the seed production areas of chili pepper in Thailand, the quality of the seed lots is low along with the presence of abnormal seeds, resulting in the reduction in seed yields. Although the average maximum air temperature in Thailand is below 33 Ž, the maximum daily temperature frequently exceeds 33 Ž during seed development in many chili production areas in the country, where extreme temperatures of Received May 10, 2006 Accepted Dec. 9, 2006 * Coressponding author Sakyo-ku Kyoto , Japan nawata@kais.kyoto-u.ac.jp more than 40 Ž sometimes occur, especially in the latter half of the dry season. Hence, high temperatures during seed development may adversely affect seed quality. Like other plant organs, seeds are affected by environmental conditions. The sensitivity of seeds to environmental stress depends on the stage of development (Gibson and Paulsen, 1999; Monjardino et al., 2005; Spears et al., 1997). After physiological maturity, i.e. the end of the seed filling stage, seed germination and vigor rapidly decrease if seeds are exposed to adverse environmental conditions (Delouche, 1980; Demir and Ellis, 1992; Doijode, 2001). This decline in quality may result from high temperature, high humidity, rainfall upon drying, and from diseases or pest attacks (Copeland and Mcdonald, 1985). The effect of high temperature stress occurring prior to physiological maturity has been demonstrated in maize (Zea mays L.), and heat stress applied during the endosperm cell division phase reduced the number of endosperm cells, starch granules (Commuri and Jones, 1999), kernel size and final dry weight (Monjardino et al., 2005). Reduction in kernel dry weight was attributed to losses in starch, protein, and oil contents with reduced activities of various enzymes (Wilhelm et al., 1999). In soybean (Glycine max L.), the standard germination rate and vigor of normal seeds produced at high

2 Pagamas et al.: Effect of high temperature on chili pepper seeds 23 temperatures (higher than 33 Ž) are low, according to Spears et al. (1997). In addition, they found that the effect of high temperature on seed quality was similar regardless of whether whole plants (95 % of the pods were brown) or individual pods (seeds harvested on the first day when they reached physiological maturity) were harvested, suggesting that the adverse effect of high temperature may occur during the seed development stages and not after physiological maturity. However, the effect of high temperature after anthesis on seed development and quality has not been fully elucidated in chili pepper. The objective of the present investigation was to opened on the same day per plant were labeled and used for the experiment. For the high temperature treatment, plants were moved from control conditions to heated plastic-covered tunnels (heat-treated) until harvest. Temperatures inside the tunnels were recorded using thermocouples connected to a data-logger (CR10X, Campbell Scientific Inc., Logan, UT) to confirm the temperature distribution. Ventilation was achieved by a window-fan attached to the tunnel. Fruit thinning or artificial pollination was not applied. Heat-treated plants were watered twice daily to ensure adequate soil moisture. There were no visible stress symptoms during the heat treatment periods. determine the effect of high temperature after anthesis on 1) the development and 2) quality and chemical composition of chili pepper seeds to gain basic knowledge for improving seed production of chili pepper in Thailand. Materials and Methods Plant material Two chili pepper (Capsicum annuun L.) varieties used in the present study consisted of Shishito (Takii Co.Ltd, Japan; SST) - a favorite variety in Japanese markets, and Huay Si Thon (HST), which was selected from several local varieties in Northeast Thailand. These two varieties were selected because flower abortion did not occur conspicuously under high Fruit and seed development harvested Fruits from labeled flowers were randomly every 5 days, starting from 5 DAA (days after anthesis) until 45 DAA in HST and 55 DAA in SST. At the time of each harvest, fruit weight, relative growth rate (on a fresh weight basis, RGR) and fruit size (length and width) were recorded for at least 25 fruits per treatment. The number of seeds per fruit and the percentage of abnormal seeds (flat, dark brown color and lighter than normal seeds), were recorded from at least 25 full-red fruits per treatment. The development of the seeds was evaluated based on the determination moisture content (SMC). of the size (width), dry weight and temperature conditions (36 }3/28 }3 Ž, mean day/night temperature), according to preliminary experiments (maximum flower abortion was 25 % in both varieties). Chili pepper seeds were sterilized by high temperature (70 Ž for 24 hours), 10 % Na3PO4 (20 min), and 1 % NaC10 (10 min) prior to sowing. Seeds were sown in vermiculite on April 12, 2005 and grown under 25 Ž conditions. Seedlings with 4-5 true leaves (four weeks after sowing) were transplanted to pots 21 cm diameter filled with gsuper soil mix h(400 mg/l N, 680 mg/l P, 540 mg/l K, plus trace elements; Akimoto Co.Ltd., Japan). Plants were grown in a plastic house at Kyoto University, Kyoto, Japan, under natural light and seasonal temperature, watered as needed and fertilized weekly with 1 L 0.1 % ghyponex h(60 g/l N, 100 g/l P, 50 g/l K; Hyponex Japan Co.Ltd., Japan). Open flowers were pinched until the onset of experiments. Seed germination and vigor For each treatment, labeled fruits were harvested at the full-red stage and kept under dry conditions at room temperature (25 Ž and 50 % relative humidity) for one week to enable slightly immature fruits to fully mature. Seeds were extracted by hand and air-dried. The seeds from each treatment were separated into normal and abnormal ones (seeds that were flat and lighter than normal seeds with a dark brown color), and only normal seeds were used for the seed germination and vigor tests. The number of seeds in each category was determined. Standard germination rate was determined using four 100-seed samples planted on 2 layers of sterilized filter paper and kept at 20/30 Ž (8 hours at 30 Ž and 16 hours at 20 Ž) for 14 days (ISTA, 2003). Seed vigor Temperature treatment was determined based on the accelerated aging germination test [2 g of seed aged at 41 Ž and nearly Among 20 pepper plants from each variety that had developed at least six reproductive nodes (approximately 11 weeks after sowing), at least 35 flowers that 100 % relative humidity for 72 hours, followed by standard germination test (Department of Horticulture and Crop Science, The Ohio State University, 2004)].

3 24 Jpn. J. Trop. Agr. 51(1) 2007 The germination treatment as follows: / GI= TiNi S index (GI) was calculated for each where Ti is the time interval (in days) between seed imbibition and germination, N; is the number of seeds that germinated on day i, and S is the total number of seeds that germinated by the end of the experiment. A low GI value indicated faster germination. Seed chemical composition Seeds from each developing fruit were extracted immediately after harvest. All the seeds were frozen in after anthesis to the naked eye. Pepper fruits that developed under high temperature conditions showed conspicuous differences (Fig. 2). Initially, the fruit weight of both varieties increased rather slowly, but the RGR was high during the 10 DAA period. Then after, the RGR decreased steadily and the fruit weight continued to increase. In HST, the maximum fruit weight appeared at around 40 DAA in both control and heat-treated fruits (Fig. 2). In SST, fruit weight followed a two-step increase, the first step, from anthesis to 15 DAA. After a transient stationary phase (15-30 DAA), the fruit weight rapidly increased again (Fig. 2). The maximum fruit weight was measured at 50 DAA and 55 DAA in the control and heat-treated fruits, respectively liquid nitrogen and stored at -40 Ž before analysis. Starch was extracted from frozen seeds and the amount was determined according to the method recommended by the Association of Official Analytical Chemists (AOAC,1995). The starch concentration was estimated using D-glucose as a standard. Protein was extracted from 5 mg of mature seeds, as described by Naito et al. (1988). The protein concentration was determined by the Bradford method with bovine serum albumin as a standard (Bradford, 1976). Fig. 1 Mean day and night temperatures during the development of pepper seeds. Solid lines indicate the mean day temperature and dotted lines indicate the mean night temperature. Thick and thin lines indicate the mean temperature for the heat stress and control conditions, respectively. Total lipid content of mature seeds was determined according to the method of Murphy and Cummins (1989) after extraction. Experimental design and statistical analysis This experiment was conducted using a completely randomized design with 2 replications of 10 plants each for fruit weight and RGR, and 3 replications for seed development, quality and chemical composition. The treatment effects were analyzed using SAS statistical analysis package (version 8.1; SAS Institute, Cary, NC) and means were compared by Tukey's studentized range test at 5 and 1 % levels of probability. Results Fruit and seed development The temperature regimes during the pepper fruit development period were 29 }2/24 } 2 Ž and 36 } 2/27 } 2 Ž (mean day/night temperature) under the control and heat stress tunnel conditions, respectively (Fig. 1). The heat-treated tunnels provided higher temperatures than control conditions. Small temperature gradients were recorded ( 0.8 Ž) inside the tunnels (data not shown). The SST and HST fruits were visible within 5 days Fig. 2 Changes in fruit weight during the development of pepper fruits. : control; : heat stress. Vertical bars show standard errors. Broken lines indicate the relative growth rate (RGR) on a fresh weight basis).

4 Pagamas et al.: Effect of high temperature on chili pepper seeds 25 (Fig. 2). Fruits of both varieties developing under control conditions displayed the maximum value of weight, which was significantly higher than that of the fruits from heat-treated increase plants (Fig. 2). However, the in the fruit length and fruit width in both varieties was not significantly different between control and heat-treated the plants (data not shown). In addition, no visible damage of fruits by the heat treatment was observed. Developing seeds were small until 10 days after anthesis, when their length was 2.5 and 1.8 mm in SST and HST, respectively. The effect of high temperature after anthesis on seed development, including SMC and dry weight increase is shown in Fig. 3. SMC of both varieties steadily decreased after anthesis, and the seeds produced under high temperature conditions, showed significantly higher SMC values than the control seeds during most of the maturation period (Fig. 3A, D). In the seeds of the control plants in both varieties, the dry weight slightly increased during the first three-week period after anthesis, and then a rapid increase occurred, that was significantly higher than that of the seeds of the heat-treated plants after 30 DAA and 35 DAA in HST and SST, respectively (Fig. 3B, E). Seeds reached harvest maturity at 45 DAA and 55 DAA in HST and SST, respectively, with the highest dry weight increase and full-red fruits. Seed width continued to increase until maturity and the seeds of the control plants were slightly longer than those of the heat-treated plants (Fig. 3C, F). Seed set was more responsive to high temperature exposure in SST than in HST. In HST, increased temperature was not associated with a significant effect on the mean number of seeds per fruit, while in SST, the mean number of seed sets was significantly reduced by high temperature treatment (Table 1). High temperature also induced the abnormal development of seeds in both varieties (Table 1). These seeds showed a complete lack of germinability (data not shown). Fig. 3 Changes in seed moisture content (SMC), dry weight, and seed width during the development of pepper fruits. : control; : heat stress. Vertical bars show the standard error. Table 1 Effect of high temperature during the seed filling stage on the seed set and abnormal seed percentage of two chili pepper varieties. * Within columns, different letters indicate a significant difference at 1 % level by Tukey Test. ns = non significant difference at 5 % level by Tukey Test. Seed germination and vigor Standard germination rate of the seeds from the control plants in both varieties was significantly higher than that of the seeds from the heat treated plants. The reduction was 28 % for HST and 25 % for SST, respectively (Table 2). Table 2 Effect of plant exposure to high temperature during the seed filling stage on seed standard germination rate and seed vigor (accelerated aging germination rate and germination index) in two chili pepper varieties * Within columns, different letters show a significant difference at 1 % level by Tukey Test.

5 26 Jpn. J. Trop. Agr. 51(1) 2007 Seed vigor was determined using the accelerated aging germination and germination index tests (Table 2). Accelerated aging germination rate of both varieties was high for the seeds of the control plants. High temperature during the seed filling stage reduced the accelerated aging germination rate in both varieties similarly. The germination index was lower in the seeds from the control plants than in those of the heattreated plants in both varieties that germination (Table 2), indicating of the seeds from the control plants was faster than that of the seeds from the heat-treated plants. Characterization of storage products in developing seeds The accumulation of storage products in both varieties was investigated in the seeds harvested from the plants in each treatment. Fig. 4 shows the changes in the total carbohydrate, protein and lipid contents in the HST and SST seeds of the control and heat-treated plants. During the early seed filling stage, the protein content of the seeds from the plants subjected to each treatment reached a maximum value and subsequently decreased. The seeds of the HST heat-treated plants showed a significantly lower protein content during the first 3-week period after anthesis, and thereafter, slightly higher content until fruit ripening than the seeds of the control plants (Fig. 4A). On the other hand, the seeds of the SST heat-treated plants showed a significantly higher protein content than the control seeds at every stage of development (Fig. 4D). In the a seeds of the control plants, the accumulation of carbohydrates started after anthesis and increased rapidly after 20 DAA and 30 DAA in HST and SST, respectively, with the highest values recorded at the seed maturity stage (Fig. 4B, E). Total carbohydrate levels of the seeds of the HST heat treated plants were significantly higher at 10, 15, and 20 DAA and then became lower than those of the seeds of the control plants until ripening (Fig. 4B). In the case of SST, the seeds of the control plants showed a significantly higher carbohydrate content than the seeds of the heattreated plants during the study period (Fig. 4E). The total lipid content in each treatment decreased slightly as the number of days after anthesis increased until 25 DAA. During this period, the lipid level of the seeds of the plants grown under different conditions in each variety was not appreciably different. Then after, the content of total lipids of the seeds of the control plants rapidly increased, while that of the seeds of the heattreated plants remained low (Fig. 4C, F). Discussion Although high temperature (36 }2/27 }2 Ž) immediately after anthesis until 5 DAA did not affect fruit growth and development, after this period, fruit development was significantly reduced by high temperature in both varieties, as indicated by the lower value of fruit weight in the heat-treated fruits than that in the control ones (Fig. 2). Bakker (1989) and Khah and Passam (1992) revealed that exposure to a high ambient temperature from the time of planting until the start of harvest induced a gradual reduction in fruit growth in sweet pepper. However, high temperature did not induce fruit malformation or visible injuries, indicating that heat affected only the quantitative development of chili pepper fruits. In SST two steps of fruit weight increase with a stationary period were observed, which is in agreement with the kinetics of seed dry weight increase (Fig. 3E). Heat stress decreased the number of seeds per fruit slightly in HST and severely in SST. Similar results were also observed in bell pepper (Khah and Passam, 1992), summer brassica (Morrison and Stewart, 2002), flax (Cross et al., 2003) and wheat (Gibson and Paulsen, 1999; Tahir and Nakata, 2005). The reduction in the number of seeds per fruit is considered to be partly responsible for the reduction in fruit development. Fig. 4 Changes in protein, carbohydrate and lipid contents during the development of pepper fruits. : control; : heat stress. Vertical bars show the standard error. High temperature during the flowering period inhibited the development of pollen grains of bell pepper and the reduction in pollen viability effectively reduced the

6 Pagamas et al.: Effect of high temperature on chili pepper seeds 27 fertilization capacity, hence the reduction in the fruit size (Erickson and Markhart, 2002) and number of seeds per fruit (Aloni et al., 2001). In this experiment, exposure to high temperatures after pollination resulted in the reduction in the seed set and fruit weight. In general, after fertilization, the development of seeds or fertilized ovules is associated with both cell division and cell enlargement. Cell division is completed during the early stages of seed development, followed by a rapid increase in fresh and dry weight due to cell enlargement. Abortion of seeds during the initial stages is probably caused by failure in proper cell division (Jones et al., 1985), which may have been induced by high temperatures. It is likely that HST which grows in Northeast Thailand under hot climatic conditions, is more tolerant to heat effects at such early seed development stages than SST, which grows in temperate Japan. Both varieties produced more than 20 % of abnormal seeds at high temperatures at high temperatures, (Table 1). In flax, there was a 3 time increase in the percentage of malformed and sterile seeds after 14 days of heat stress (Cross et al., 2003). Abnormal development of seeds apparently reduces the ability of seeds to germinate and to produce normal and vigorous seedlings (Spear et al., 1997). The high proportion of abnormal seeds produced under high temperature conditions (Table 1) is likely to reduce the quality of seed lots and may adversely affect fruit development. Although the cause of the increase in the proportion of abnormal seeds induced by high temperature has not been elucidated, the direct effect of high temperature may have affected seed development. Following the translocation of photosynthates to the endosperm and embryo, sugars, amino acids and minerals are resynthesized into specific storage compounds, which can be generally categorized into carbohydrates, oil, proteins and phytin (Bewley and Black, 1985). The accumulation of storage compounds is markedly diversified among species, e.g. lipids and proteins in oilseed rape (Paulo et al., 1997; Eastmond and Rawsthorne, 2000), carbohydrates in cereals and grasses, and carbohydrates, with lesser amounts of proteins in pulses (Copeland and McDonald, 1985). The present results showed that chili pepper seeds accumulated carbohydrates and lipids as major storage products with small amounts of proteins at maturity (Fig. 4). Seed storage accumulation is known to be affected by heat stress (Wilhelm et al., 1999; Monjardino et al., 2005; Thomas et al., 2003; Tahir and Nakata, 2005). In the present study, heat stress after anthesis strongly reduced carbohydrate and lipid accumulation in chili pepper seeds, especially during the maturation period (Fig. 4). The major storage lipid of seeds is in the form of triacylglycerols (Bewley and Black, 1985), whose synthesis involves various cellular components, among which sucrose is the first substrate in the biochemical pathways related to the conversion of carbohydrates to fatty acid (Bewley and Black, 1978; Bewley and Black, 1985). In the developing seeds, sucrose from the mother plants is the initial source of substrates starch and lipid synthesis. Low carbohydrate for or lipid levels in the seeds of the heat-treated plants suggest that the ability of assimilation or translocation into seed was limited (Thomas et al., 2003; Monjardino et al., 2005; Wilhelm et al., 1999). In general, elevated temperatures increase the respiration rate and inhibit the photosynthesis metabolism (Alscher and Cumming, 1990). High temperature during the grain filling period inhibited photosynthetic processes in wheat (Al-Khatib and Paulsen,1990) and reduced N remobilization from stem and tiller to seed (Tahir and Nakata, 2005). Thus heat stress may have reduced the supply of photosynthates and other nutrients for seed development from current assimilation, resulting in the reduction in lipid and carbohydrate accumulation, which may be a major factor for the reduction in seed dry weight. Seed quality is maximal at the end of the seed filling stage, and viability and vigor begin to decrease immediately thereafter (Demir and Ellis,1992). Reduction in lipid and carbohydrate accumulation through the suppression of photosynthates and other nutrients may also affect the seed quality. In general, the supply of assimilates exerts a major effect on the number of fertilized embryos that continue to develop. Thus, it is possible that heat stress after anthesis is particularly detrimental to subsequent seed development, due to the disturbance in carbohydrate and lipid accumulation. As a result, the standard germination rate and vigor (accelerated aging germination rate and germination index) of the chili pepper seeds produced under high temperature conditions were considerably reduced (Table 2). Keigley and Mullan (1989), Spears et al. (1997), and Zankis et al. (1994) also reported that increase of the temperature during the seed development and maturation stages reduced soybean seed quality. In the present study, although the HST and SST plants grew well under high temperature conditions without visible symptom of injury, seed quality was

7 28 Jpn. J. Trop. Agr. 51(1) 2007 adversely affected. This suggests that high temperature after anthesis affected seed development and quality directly, and not through the growth of the whole plant. Moreover, high temperature in our experiment was maintained for approximately 8 hours each day after anthesis until harvest, whereas in fields, the temperature is usually high for only a few hours each day. Thus the effect of high temperature in our experiment may have been overestimated compared to that observed in the plants growing in ordinary fields. Cross et al. (2003) demonstrated that higher percentages of abnormal and ungerminable seeds were produced during a longer period of heat stress. Additional studies should be carried out to determine the direct effect and critical duration of high temperature exposure affecting chili pepper seed quality. References Al-Khatib, K. and G. M. Paulsen Photosynthesis and productivity during high-temperature stress of wheat genotypes from major world regions. Crop Sci. 30: Aloni, B., M. Peet, M. Phart and L. Karni The effect of high temperature and high atmospheric CO2 on carbohydrate change in bell pepper (Capsicum annuum) pollen in relation to its germination. Physiol. Plant. 112: Alscher, R. G. and J. R. Cumming Stress responses in plants: adaptation and acclimation mechanism. Wiley-Liss, New York, p407. Association of Official Analytical Chemists Official methods of analysis, l6thedn. AOAC, Virginia, USA. Barker, J. C The effect of temperature on flowering, fruit set and fruit development of glasshouse sweet pepper (Capsicum annuum L.). J. Hurt. Sci. 64: Bewley, J. D. and M.Black Physiology and biochemistry of seeds. Berlin Heidelberg, New York, p306. Bewley J. D. and M. Black Seeds: Physiology of development and germination. Plenum Press, New York, p367. Bradford, M. M A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: Commuri, P. D, and R. J. Jones Ultrastructural characterization on maize (Zea mays L.) exposed to high temperature during endosperm cell division. Plant Cell Environ. 22: Copeland, L. O. and M. B. McDonald Principle of seed science and technology. Burgess Publishing. USA, p321. Cross, R. H., S. A. B. Mckay, A. G. Mchughen and P. C. Bonham- Smith Heat stress effects on reproduction and seed set in Linum usitatissimum L. (flax). Plant Cell Envir. 26: Delouche, J. C Environmental effects on seed development and seed quality. HortSci.15: Demir, I. and R. H. Ellis Development of pepper (Capsicum annuum) seed quality. Annal. Appl. Biol. 121: Department of Horticulture and Crop Science, Ohio State University Seed vigor and vigor tests. Available at `seedsci/svvt03.html. Doijode, S. D Seed storage of horticultural crops. Food Prod. Press, New York, p339. Eastmond, P. J. and S. Rawsthorne Coordinate changes in carbon partitioning and plastidial metabolism during the development of oilseed rape embryos. Plant Physiol. 122: Erickson, A. N. and A. H. Markhart Flower developmental stage and organ sensitivity of bell pepper (Capsicum annuum L.) to elevated temperature. Plant Cell Envir. 25: Gibson, L. R. and G. M. Paulsen Yield components of wheat grown under high temperature stress during reproductive growth. Crop Sci. 39: International Seed Testing Association International rules for seed testing. International Seed Testing Association, Switzerland, p327. Jones, R. J., J. Roessler and S. Ouattar Thermal environment during cell division in maize: Effect on number of endosperm cells and starch granules. Crop Sci. 25: Keigley, P. J. and R. E. Mullan Change in soybean seed quality from high temperature during seed fill and maturation. Crop Sci. 26: Khah, E. M. and H. C. 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Nakata Remobilization of nitrogen and carbohydrate from stem of bread wheat in response to heat stress during grain filling. J. Agron. Crop Sci. 191: Thomas, J. M. G., K. J. Boote, L. H. Allen, Jr., M. Gallo-Meagher and J. M. Davis Elevated temperature and carbon dioxide effects on soybean seed composition and transcript abundance. Crop Sci. 43: Wilhelm, E. P., R. E. Mullen, P. L. Keeling and G. W. Singletary Heat stress during grain filling in Maize: Effect on kernel growth and metabolism. Crop Sci. 39: Zankis, G. N., R. H. Ellis and R. J. Summerfield A comparison of changes in vigour among three genotypes of soybean (Glycine max) during seed development and maturation in three temperature regimes. Exp. Agric. 30:

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