J. Sci. Dev. 2009, 7 (Eng.Iss.1): 1 - 8 HA NOI UNIVERSITY OF AGRICULTURE 1 Photosynthetic and root characters related to drought tolerance in plant Các đặc tính quang hợp và rễ liên quan đến chịu hạn ở cây lúa Pham Van Cuong Department of Food Crop Science, Faculty of Agronomy, Hanoi University of Agriculture TÓM TẮT Thí nghiệm trong chậu được tiến hành nhằm đánh giá các đặc tính sinh lý và đặc tính rễ liên quan đến tính chịu hạn của các giống lúa bao gồm lúa nước loài phụ indica, lúa nước loài phụ Japonica, lúa lai từ dòng bất dục đực nhân mẫn cảm nhiệt độ và loài lúa dại để so sánh với giống lúa Indica chịu hạn. Ở giai đoạn đẻ nhánh hữu hiệu, các chậu thí nghiệm được rút cạn nước rồi để khô trong vòng 4 ngày, sau đó được tưới nước trở lại. Ở giai đoạn trước rút nước, hạn và sau phục hồi (sau tưới nước 4 ngày), chọn ngẫu nhiên 4 chậu của mỗi giống để đo các chỉ tiêu về quang hợp như cường độ quang hợp, độ dẫn khí khổng, cường độ thoái hơi nước và chỉ số khối lượng riêng của lá (một chỉ tiêu đánh giá độ dày lá). Những cây đo quang hợp được chọn để đo độ dẫn nước và các đặc tính về rễ như chiều dài rễ tối đa, số lượng rễ và khối lượng rễ khô. Khi xử lý hạn hầu hết các đặc tính quang hợp như cường độ trao đổi CO 2 , độ dẫn khí khổng và cường độ thoát hơi nước đều giảm mạnh ở lúa lai F1 và lúa dại tương tự như ở giống lúa chịu hạn (CH5), trong khi đó những chỉ tiêu này giảm ít hơn ở các giống lúa khác. Ngược lại những chỉ tiêu này lại phục hồi tốt hơn ở lúa lai F1 lúa dại và giống lúa chịu hạn so với ở các giống khác. Cường độ quang hợp tương quan thuận với độ dẫn khí khổng và cường độ thoát hơi nước ở tất cả các giống lúa ở giai đoạn hạn và phục hồi, tuy nhiên tương quan này không ở mức ý nghĩa tại giai đoạn trước rút hạn. Cường độ quang hợp và chỉ số diệp lục (SPAD) có tương quan thuận ở giai đoạn trước rút hạn và sau phục hồi, tuy nhiên tương quan này lại âm ở giai đoạn hạn. Cường độ quang hợp có tương quan thuận với số lượng rế/cây ở giai đoạn phục hồi. Từ khoá: Cây lúa, chịu hạn, kiểu gien, quang hợp, rễ. SUMMARY The pot experiment was conducted to estimate physiological and root characters related to drought tolerance of different cultivated rice cultivars including low land Indica subspecies, low land Japonica subspecies and F1 hybrid from thermo-sensitive genic male sterile line and wild rice specie (Oryza Rufipogon) in compared with drought tolerance cultivar (Indica low land rice). At the active tillering stage, the plants were moved out all waters the kept dried during for four days, after that the pots were recovering. Four plant of each cultivars was randomly selected for measuring photosynthetic characters viz., photosynthetic rate, stomatal conductance, transpiration rate and specific leaf area (SLA a revert indicator of leaf thickness). The plant measured photosynthetic characters were sampled for measuring water conductance and root characters such as root length, number of root per plant and root dry weight. As drought treatment, most photosynthetic character such as CO 2 exchange rate, stomatal conductance and transpiration rate decreased in F1 hybrid and wild rice as much as in drought tolerance cultivar (CH5), whereas they were much higher in the other cultivars. In contrary, these characters recovered in all F1 hybrid, wild rice and check cultivar were quite higher than those in the others at recovering stage. Photosynthetic rate were significantly and positively correlated with stomatal conductance and transpiration rate in all rice cultivars at both drought and recovering stages, whereas the correlation was not significant at before drought stage. A correlation between CER and SPAD was observed to be positive at before drought and recovering stages, however it was negative at drought stage. CER was positively correlated with the number of root per plant at recovering stage. Journal of Science and Development 2008: Tập VI, No 6: 179- HA NOI UNIVERSITY OF AGRICULTURE 2 Key words: Drought tolerance, photosynthesis, rice plant, root. 1. INTRODUCTION Water is essential to plant growth because it provides the medium within which most cellular functions take place. Nowadays, the restricted availability of water resource has focused attention. On paddy and upland fields where the irrigation water depends mainly on rainfall, rice plants are often exposed to drought (O'Toole, 1982). On the other hand, about 70% of the rice cultivation area in Southeast and South Asia, which is about half of the total world production, depends on irrigation from rainfall. From the worldwide viewpoint, the fact indicates that studies on drought resistance are very important at present. According to mechanisms of drought resistance, the responses to drought in plants relate to photosynthetic rate, stomatal conductance, transpiration rate, leaf rolling, enhanced root growth, specific leaf area and so on. Accumulated evidence suggests that both chemical and hydraulic signal are operative and integrated in the regulation of leaf growth and stomatal conductance when plants grown under drought stress (Davies et al., 1994; Comstock, 2002). The hydraulic signals involve changes in pressure potential in the xylem and changes in water content of the guard cells and other epidermal cells. The chemical signals relate to abscisic acid (ABA), which is synthesized in roots under water stress and transported to leaves and/or together with ABA synthesized in the leaves themselves, induce stomatal closure. Also, root development is fundamentally involved in the response to many plant stresses, such as drought (Adam et al., 2002). The possession of a deep and thick root system which allows access to water deep in the soil profile is considered crucially important in determining drought resistance in upland rice and substantial genetic variation exists for this (Fukai and Cooper, 1995; O'Toole, 1982; Yoshida and Hasegawa, 1982). The drought resistance of cultivated plants, including rice plants, refers not only to their specific ability, such as survival capacity or subsequent recovering ability, but also to their integrated abilities in the production processes through their growing period (Fukai and Cooper, 1995; Mambani and Lal, 1983). In this study, we estimated the drought tolerance in F 1 hybrid, wild rice, upland rice and low land rice cultivars in compared with drought tolerance rice at the active tillering stage by measuring the photosynthetic rate, stomatal conductance, water conductance and root characters. 2. MATERIALS AND METHODS 2.1. Plant Materials The experiments were carried out with 8 different rice cultivars: CH5 (Drought resistance used as check cultivar), Khau Suu (local upland rice cultivar), 103S (low land, TGMS line -indica), R20 (low land, restorer line- indica), Oryza Rufipogon (wild rice species), Vietlai 20 (F 1 hybrid rice - 103S/R20), Lily 328 (low land rice, susceptible for drought, japonica): and Toitsu (low land rice, susceptible for drought, japonica). The experiments were set in green house in Faculty of Agronomy, Hanoi University for Agriculture, Vietnam during spring cropping season in 2006. 2.2. Planting Seed of rice cultivars were incubated and sown in the seedling bed (60 x 35 x 8 cm). Seedling at the 3 - 4- leaf stage was transplanted singly into Wager pot (0.02 m 2 ), one seedling per one pot (Gomez and Gomez, 1984). 2.3. Fertilization Total fertilizer was applied with N, P 2 O 5 and K 2 O was at the rate of 0.48, 0.36 and 0.36 (g per pot), respectively. Basal dressing for one pot with N, P 2 O 5 and K 2 O was at the rate of 0.16, 0.18 and 0.12 g, respectively. Top dressing at 7 DAT and 14 DAT with N, P 2 O 5 and K 2 O at the rate of 0.08, 0.18 and 0.08 g, respectively. Final dressing at the panicle initiation stage (20-18 days before heading) was applied with N and K 2 O at the same rate of 0.06 g. 2.4. Drought treatment At the active tillering stage (30 days after transplanting), all the water inside in the pots were moved out then the pots were kept dried during 4 days, after that re-watering (O'Toole, 1982; Fukai and Cooper, 1995). 2.5. Measurement Photosynthetic parameters: At three stages as before draining, during drought treatment and 4 days after recovering, 4 plants of each rice cultivars were randomly selected for measuring photosynthetic Pham Van Cuong 3 characters. Two top-fully expanded leaf of each plant was selected for measuring CO 2 exchange rate (CER), transpiration rate (Tr) and stomatal conductance (gs) using LICOR 6400, at temperature of 30 o C, light intensity of 1500 mol/m 2 /s and CO 2 concentration of 370 ppm. The plants measured photosynthesis were selected for measuring SPAD value (an indicator of chlorophyll content) using SPAD, Motorola, 502, Japan, Specific leaf area (SLA), water conductance and root characters: The leaf measureed CER was sampled for measuring leaf area using ANA-GA-5, Japan, then dried at 80 o C for 48h for calculating SLA. SLA = Leaf area/ Leaf dry weight Water conductance: The plants were cut at 5 cm high from the base of plant and use dry cotton to cover the stem. After two hours the cotton was weighted for calculating water conductance. Root characters: The plants were selected maximum root length and counted the number of active root per plant. The roots of each plant were dried at 80 o C for 48 hours for root dried weight measurement. Statistical analysis: Data was analyzed by SAS program Ver.8.2, (1990) after Shinjo (1994)[f1]. Means were tested by least significant difference at P0.05 level . At each stage, correlation among photosynthesis and root characters were calculated of 4 plants considered as 4 replications. 3. RESULTS AND DISCUSSION CO 2 Exchange Rate: Table 1 showed that the CO 2 exchange rate (CER) of all 8 cultivars decreased under drought condition (measured 4 days after draining) and increased in recovering stage (4 days after watering). The CER decreased in wild rice (Oryza Rufipogon; from 31.23 molm - 2 s -1 to 18.46 molm -2 s -1 ) and F1 hybrid (Vietlai 20; from 30.47 molm -2 s -1 to 20.18 molm -2 s -1 ) as much as in check cultivar (CH5; from 32.36 molm -2 s -1 to 15.65 molm -2 s -1 ), but it was higher than that in the others cultivars. After recovering, the CER value in both CH5 (31.32 molm -2 s -1 ) and F1 hybrid (28.74 molm -2 s -1 ) were almost the same as it before draining. However at recovering stage, the CER of Oryza Rufipogon was lower than that of other cultivars, and lower than that at before draining stage. Stomatal Conductance (Gs): The stomatal conductance of all rice cultivars reduced after draining and increased after recovering (Table 2). The Gs of CH5, Oryza Rufipogon and Vietlai 20 dropped significantly during drought treatment (especially with CH5, Oryza Rufipogon) and recovered to the same level as of the well-watered treatment (with CH5 and Vietlai 20). This may be the changes in pressure potential in the xylem and changes in water content of the guard cells and other epidermal cells (Davies et al.,1994; Comstock, 2002). CER was not correlated with and stomatal conductance in all rice plant at before draining stage (r = 0.28), whereas it was significant at drought at (r = 0.90) and recovering stage (r = 0.59). This might be due to abscisic acid (ABA), which is synthesized in roots under water stress and transported to leaves and induce stomatal closure (Davies et al .,1994). Table 1. CO 2 exchange rate of rice cultivar at the active tillering stage (mol m -2 s -1 ) Cultivars Before draining 4 days after draining 4 days after recovering 103S /R20 30.47 20.18 28.74 R20 32.09 24.23 29.81 103S 33.22 27.7 28.93 CH5 (check) 32.36 15.65 31.32 Khau Suu 28.28 22.62 25.88 Oryza Rufipogon 31.23 18.46 24.97 Photosynthetic and root characters related to drought tolerance in plant 4 Lily 328 32.75 26.85 30.64 Toitsu 32.20 22.33 31.12 LSD5% 1.31 1.87 1.07 Table 2 . Stomatal conductance of rice cultivar at the active tillering stage (mol m -2 s -1 ) Cultivars Before draining 4 days after draining 4 days after recovering 103 S /R20 0.70 0.10 0.80 R20 0.84 0.26 0.80 103 S 0.65 0.32 0.61 CH5 (check) 0.84 0.03 0.81 Khau Suu 0.52 0.26 0.67 Oryza Rufipogon 0.49 0.04 0.29 Lily 328 0.45 0.30 0.56 Toitsu 0.70 0.14 0.90 LSD5% 0.13 0.05 0.12 Fig. 1. Correlation between CO 2 exchange rate (CER) and stomatal conductance (gs) in rice cultivars before draining (A), 4 days after draining (B) and 4 days after recovering stages (C) B y = 28.83x + 17.21 r = 0.90*** 15 17 19 21 23 25 27 29 0.0 0.2 0.4 0.6 0.8 1.0 CER ( molm -2 s-1 C y = 8.60x + 23.02 r = 0.59* 23 25 27 29 31 33 0.0 0.2 0.4 0.6 0.8 1.0 Gs(molm -2 s -1 ) Ay = 2.7202x + 29.754 r = 0.26 27 29 31 33 35 0.0 0.2 0.4 0.6 0.8 1.0 1.2 1.4 1.6 O.Rufipo gon CH5 Vietlai20 R20 Khausuu 103s Lily Toitsu Pham Van Cuong 5 Transpiration rate: Similar to the changes of photosynthetic rate and stomatal conductance, transpiration rate of all rice cultivars decreased after draining water and recovered almost similar to the same of that after recovering (Table 3). As drought treatment, transpiration rate reduced significantly and strongly in CH5, following in Vietlai 20 and Oryza Rufipogon. The reduction was less in the other cultivars than that in check variety. The reduced in transpiration rate cause to reduce the effect of water deficit, the early drought adaptation of plants. Thus, both the F1 hybrid and wild rice could be good for drought tolerance. At recovering time, the transpiration rate increased to the same of that before with CH5, little bit higher than before with Vietlai 20, Toisu , Khau Suu and Lily 328. A significant and positive correlation was observed between CO 2 exchange rate and transpiration rate during drought condition (r = 0.89) and recovering time (r = 0.78), whereas it was not significant at the time before draining (r = 0.31) (Fig.2). At the drought time, which cultivars had transpiration rate reduced much, that cultivars would had low CER (CH5, F1 hybrid and wild rice). At the recovering time, the transpiration rate of CH5 and F1 hybrid were similar before and the CER could recover well. The lower transpiration rate in F1 hybrid and wild rice which is similar to check cultivars was caused by better stomatal system. Table 3 . Transpiration rate of rice cultivar at the active tillering stage (mmol m -2 s -1 ) Cultivars Before draining 4 days after draining 4 days after Recovering 103 S /R20 8.96 0.39 10.26 R20 10.55 4.38 9.71 103 S 9.94 4.53 8.88 CH5 (check) 11.82 0.53 11.15 Khau Suu 8.44 4.00 9.48 Oryza Rufipogon 8.24 1.02 5.34 Lily 328 8.04 6.09 9.34 Toitsu 7.94 3.89 11.31 LSD5% 1.06 0.89 1.35 A y = 0.35x + 28.26 r = 0.31 27 29 31 33 35 0 2 4 6 8 10 12 14 B y = 1.70x + 16.97 r = 0.89** 15 20 25 30 0 2 4 6 8 10 12 14 CER(mmolm -2 s -1 ) C y = 0.99x + 19.54 r = 0.78** 23 25 27 29 31 33 0 2 4 6 8 10 12 Tr (mmol m -2 s -1 ) Fig. 2. Correlation between CO 2 exchange rate (CER) and transpiration rate (Tr) in rice cultivars before draining (A), 4 days after draining (B) and 4 days after recovering stages (C). Note: Symbols were the same as shown in Fig.1 Photosynthetic and root characters related to drought tolerance in plant 6 SPAD value: There were non-significant differences in SPAD values in all rice cultivars during three stages of experiment (Table 4). At before draining and drought stage, the SPAD of each cultivars were almost the same each other. At recovering stage, the SPAD decreased in Khau Suu, Oryza Rufipogon, 103S, Vietlai 20 and CH5. Among of them, the decrease was much in Khau Suu, following in 103S, Vietlai 20, Oryza Rufipogon and CH5. Rice plants grown under water deficit, the SPAD reduced with cultivars which were considered to be drought resistance. The correlation between CER and SPAD was shown in Fig.3. CER was positively correlated with SPAD value at both before draining stage (r = 0.41) and recovering stage (r = 0.57), but the correlation was not significant at drought stage (r = -0.34) Table 4. SPAD value of rice cultivars at the active tillering stage Cultivars Before draining 4 days after draining 4 days after recovering 103 S /R20 38.39 39.30 36.04 R20 40.83 39.58 41.33 103 S 36.87 37.65 33.99 CH5 (check) 39.08 40.49 38.63 Khau Suu 36.20 35.46 31.16 Oryza Rufipogon 39.20 38.85 37.50 Lily 328 38.97 39.51 41.27 Toitsu 37.89 38.91 37.94 LSD5% 1.75 2.30 2.45 A y = 0.44x + 14.38 r = 0.41* 27 29 31 33 35 30 32 34 36 38 40 42 B y = -0.90x + 57.48 r = -0.34 15 17 19 21 23 25 27 29 30 32 34 36 38 40 42 CER (mmolm -2 s -1 ) C y = 0.39x + 14.34 r = 0.57* 20 22 24 26 28 30 32 30 32 34 36 38 40 42 SPAD Pham Van Cuong 7 Fig. 3. Correlation between CO 2 exchange rate (CER) and SPAD in rice cultivars before draining (A), 4 days after draining (B) and 4 days after recovering stages (C). Note: Symbols were the same as shown in Fig.1. Specific leaf area (SLA): Vietlai 20 has the highest value of SLA, following are 103S, R20, Toisu, Lily 328, Khau Suu, CH5 and Oryza Rufipogon has the lowest value of SLA (Table 5). Water evaporation through leaf is a driving force of water flow from root to shoot. When water evaporation is low, root pressure plays an important role to maintain the water flow from root to shoot. The specific leaf area of plants is one of the genotype that related to genetic. Many observations pointed that cultivars that have thin leaves (the value of SLA is high) that cultivars are considered to be drought resistance. Because of thin blade, leaf will be easy to roll up to reduce CO 2 exchange rate, stomatal conductance and transpiration rate. CH5 and Oryza Rifipogon had smallest value of SLA. Root characters: The cultivar that had the longest maximum root length is CH5, following are KhauSuu, Oryza Rufipogon, Toitsu, Vietlai 20, Lily 328, R20 and 103S. It means CH5 and Oryza Rufipogon had deeper roots (Table 5), which increasing the ability to extract soil water from depth (Mambani and Lal, 1983a,b,c; Yoshida and Hasegawa, 1982). Despite the longest root length, the number of root per plant was much higher in F1 hybrid (292) than that in CH5 (211) and Oryza Rufipogon (232), indicating that the F1 hybrid had thick root system, which also allows access to water deep in the soil profile is considered crucially important in determining drought resistance in upland rice and substantial genetic variation exists for this (Ekanayake et al., 1985b; Fukai and Cooper, 1995; O'Toole,1982; Yoshida and Hasegawa, 1982). A significant and positive correlation was observed between CER and the number of root per plant in all rice cultivars at recovering stage, indicating that the greatest thick root system in F1 hybrid not only contributed to resistance during drought condition but also at recovering stage. Water conductance: Water conductance from root to stem was shown in Table 5. In detail, the water conductance of Vietlai 20, CH5 and Toitsu was 0.55, 0.49 and 0.46, respectively, and it was higher than that in the other cultivars. The result indicates that the cultivars had high value of water conductance, that also had high value of photosynthesis, stomatal conductance and transpiration rate at recovering stage and it contributed to drought resistance Table 5. Specific leaf area (SLA), root characters and water conductance in rice cultivars at recovering stage Cultivars SLA (cm 2 g -1 ) Maximum root length (cm) Number of root plant -1 Root dry weight (g plant -1 ) Water conductance (mg plant -1 s -1 ) 103 S /R20 259.68 24.81 292.00 7.54 0.55 R20 227.48 22.63 252.33 6.94 0.33 103 S 234.31 19.76 275.33 6.89 0.38 CH5 (check) 200.89 27.83 211.00 6.66 0.49 Khau Suu 219.99 27.75 247.33 6.03 0.40 Photosynthetic and root characters related to drought tolerance in plant 8 Oryza Rufipogon 197.29 27.70 232.00 4.17 0.29 Lily 328 223.48 24.83 283.66 3.56 0.38 Toitsu 227.52 25.00 283.00 4.84 0.46 LSD5% 21.27 1.17 17.17 0.75 0.05 Fig. 4. Correlation between CO 2 exchange rate (CER) and number of root per plant in rice cultivars at recovering stage Note: Symbols were the same as shown in Fig.1 4. CONCLUSION Among rice cultivars, F1 hybrid (Vietlai 20) and wild rice (Oryza Rufipogon) was the best for photosynthetic characters which similar to check cultivar for drought tolerance. During drought condition, low photosynthetic rate and transpiration rate caused by low stomatal conductance contributed to drought tolerance in rice plant. The larger number of root per plant was the reason for better recovering of rice plant after drought treatment. REFERENCES Adam HP, Jill EC, Peter H, Hamlyn GJ, Howard G. (2002). Linking drought-resistance mechanisms to drought avoidance in upland rice using a QTL approach: progress and new opportunities to integrate stomatal and mesophyll responses. Journal of Experimental Botany 53,989-1004. Comstock JP. (2002). Hydraulic and chemical signalling in the control of stomatal conductance and transpiration. Journal of Experimental Botany 53,195-200. Davies WJ, Tardieu F, Trejo CL. (1994). How do chemical signals work in plants that grow in drying soil? 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(1983c). Response of upland rice varieties to drought stress. III. Estimating root configuration from soil moisture data. Plant and Soil 73, 95-104. O'Toole JC. (1982). Adaptation of rice to drought- prone environments. In: Drought resistance in crops with the emphasis on rice. Manila: IRRI, 195-213. Shinjo, A. (1994). Introduction to Statistics by PC SAS. (in Japanese) Tokai Univ. Press, Hadano, pp 195. Yoshida S, Hasegawa S. (1982). The rice root system, its development and function. In: Drought resistance in crops with the emphasis on rice. Manila: IRRI, 83-96 . their growing period (Fukai and Cooper, 1995; Mambani and Lal, 1983). In this study, we estimated the drought tolerance in F 1 hybrid, wild rice, upland rice and low land rice cultivars in compared. important in determining drought resistance in upland rice and substantial genetic variation exists for this (Fukai and Cooper, 1995; O'Toole, 1982; Yoshida and Hasegawa, 1982). The drought. estimate physiological and root characters related to drought tolerance of different cultivated rice cultivars including low land Indica subspecies, low land Japonica subspecies and F1 hybrid from