文献翻译-低温胁迫下磷肥对水稻耐寒性及相关生理参数的影响.doc

低温胁迫下磷肥对水稻耐寒性及相关生理参数的影响侯立刚，陈文福，赵国臣，马巍，齐春艳，刘 亮，孙洪娇1. 沈阳农业大学水稻研究所，沈阳100866,中国2. 吉林省农科院水稻研究所,吉林，公主岭，136100，中国摘要：本文旨在研究磷肥强化水稻耐寒性的生理机制。通过试验，研究了低温胁迫下不同水平磷肥对水稻幼苗（非耐寒性品种长白9号及耐寒性品种吉粳81号）耐寒性及相关生理参数的影响，同时确认耐寒性。与常温条件比较，低温条件下水稻的相关叶绿素含量，光合速率，Fv/Fm，qP降低，及不饱和脂肪酸指数升高。与耐寒性栽培品种比，对非耐寒性品种影响更大。适当增加磷肥用量可增强水稻幼苗素质，减缓水稻幼苗相关叶绿素含量降低程度，增加光合速率，Fv/Fm，qP及不饱和脂肪酸指数，增强耐寒性栽培品种低温条件下生存能力。 施磷量增高对耐寒性栽培品种影响更大。磷酸盐调节光合生理及膜流动性，缓减低温下植株损害，增加水稻耐寒能力。关键词：低温胁迫，磷肥，水稻，耐寒性，生理参数引言地处东亚季风区域，中国是世界上气候脆弱区域之一。根据资料统计，仅农业领域我国每年因气象灾害造成的农作物受害面积就达48亿公顷，给我国粮食生产造成严重经济损失。低温冷害是我们重大农业气象灾害，我国每年稻谷损失达100-500亿（王，2008）。过去50年（1951-2000）对吉林省气象数据分析表明，每年气温都有一定涨幅，但在稻米生长的重要季节，区域性及阶段性的低温冷害仍然不时发生，且有变的更加频繁和严重的迹象，因此会形成低温冷害年（俞等人，2003）。作为植物生长与发育的必要营养成分之一，磷以多种方式参与植物生命活动。越来越多的研究表明，磷可以大大提高植物的耐性，尤其是耐寒性（王等人，2003；李等人，2004；魏等人2005，陈等人，2007）。多年实践已经证实低温 会影响水稻对磷的吸收，阻碍生长；而增加磷肥可以促进水稻根系生长，帮助移栽植株返青，因此可以提高 稻米产量。近几年来，对甘蔗（魏等人，2003），西瓜（陈等人，2007），冬小麦（李等人，2004）及烟草（王等人，1999）植被的研究表明，适量增加磷肥用量对改善植株耐寒性发挥重要作用。然而，对于增加磷肥如何提高水稻幼株耐低温性的生理机理方面的研究还不多。本研究旨在研究这一机理，借助两种耐寒性水稻品种作为研究材料，研究低温条件下增加磷肥对水稻幼株的耐寒性及相关生理特征影响。目的是为了进一步阐明生理机理，为以磷肥为中心的抗冷冻技术措施提供理论依据。 材料与方法试验材料本试验于2010年在吉林省农科院育苗基地进行，以不同耐寒品种吉粳81号（耐寒性品种）长白9号（非耐寒性品种）作为试材。试验设计：首先，选择成熟度及丰满度相同的种子，与2010年4月中旬浸泡这些种子，使其生成潜在幼芽。然后，选择幼芽生长速度相同的种子，均匀地播种到育苗盘里，（育苗盘里装有不同的磷酸盐），播种密度为每个播种盘里200 g m-2。5种磷肥施用量（P2O5）- 0，25，50，75，100 g m-2分成两组，一组接受低温处理，一组用于常温比较，每个组处理过程重复两次。最后，灌溉足量水分，参考吉林省高产量稻米培育方法覆盖在幼苗温室内。 温度处理水稻幼苗长出3片幼叶时，把经低温处理的一组移到人工气候室（LT/ACR-2002，易盛泰和公司人工气候室系统），连续5天接受低温胁迫，每日平均温度8；参考表1特定参数设置。另一组放置室外，温度数据从国际稻米研究机构天气记录获取；具体温度情况见图1（公主岭，2010）。过程中日平均温度超过15，并定期浇水。表1 气候控制室温度及光照度设置Time：时间Daily average temperature：8：每日平均温度：8Light (lx) ：光照度（lx）Temperature ()：温度()FL：荧光灯组数；NL：钠灯组数图1 自然空气温度变化Min temperature最低温度Max temperature最高温度Average temperature平均温度Temperature ()温度()衡量项目及方法水稻幼苗耐寒能力评估参考韩等人（2004）及李等人（2006）的方法，结合吉林省的实际情况，本研究通过评估叶子的赤枯度对水稻幼苗生长能力进行鉴定和评级。标准如下：1级-秧苗赤枯率20%；2级-秧苗赤枯率21%-30%；3级-秧苗赤枯率31%-40%；4级-秧苗赤枯率41%-50%；5级-秧苗赤枯率51%-60%；6级-秧苗赤枯率61%-70%；7级-秧苗赤枯率71%-80%；8级-秧苗赤枯率81%-90%；9级-秧苗赤枯率90%生物量测定之后，从每组选择100株幼苗，用蒸馏水冲洗，再105下放置15分钟，然后烘干获取这100株幼苗干重（克）。叶绿素含量测定叶绿素含量用叶绿素计(SPAD-502 型号)测定。用叶绿素计夹住完全展开叶片的上缘中心职位，测量叶绿素含量值。从一组拿10株幼苗，获取他们的平均叶绿素含量值。重复这一过程三次。净光合速率测定用美国LI公司生产的LI-6400便携式光合作用仪测量水稻叶片的净光合速率（Pn）。叶绿素荧光参数测定用德国WALZ公司PAM2500便携式脉冲调制叶绿素荧光分析仪测定水稻叶片叶绿素荧光参数。测量方法参考让蒂等人（1989）的方法。取完全展开叶片最上部，测量前将待测叶片暗适应20分钟，然后打开测量灯，获取初始荧光F0，随后打开饱和脉冲灯，得到最大荧光Fm及最大光化学效率Fv/Fm。然后打开光化学灯，待光适应20分钟后，测得光存在时最大荧光Fm等荧光参数。其中，Fv/Fm、qP、qN由下列公式计算得出：Fv/Fm（最大光学效率）=（Fm-Fn）/Fm；qP（光化学猝灭系数）=（Fm-Ft）/（Fm-Fn）；qN（非光化学荧光猝灭系数）=Fm/ Fm-1。参照俞等人（1996）方法测定叶片油酸、亚油酸和亚麻酸含量。其中，不饱和脂肪酸指数（UFA）=油酸含量（mol%） 1+亚油酸含量（mol%） 2+亚麻酸含量（mol%）3.结果低温条件下磷肥对水稻幼苗耐寒性的影响不同品种间耐寒程度存在明显差异（见表2）。低温条件下，未施用磷肥吉梗81号赤枯率为17.6%耐低温级别为1级，而长白9号赤枯率为31.2%，耐低温等级为3级。当施磷量在075 g m-2时，吉粳81号和长白9号赤枯率随着施磷量的增加而下降，其中吉粳81号变化幅度较小，耐低温级别未发生明显；而长白9号赤枯率变化明显，施磷量为75 g m-2时赤枯率比未施磷下降37.8%，耐低温级别从3及上升到1级。当施磷量达到100 g m-2时，吉粳81号和长白9号赤枯率有所上升，其中长白9号上升明显，耐低温级别由1级降到2级。表2 低温条件下磷肥对水稻幼苗耐寒性的影响Phosphorus application (g m-2) 施磷量g m-2)Jijing 81 Changbai 9吉粳81号 长白9号Seedling mortality (%) 赤枯率（%）Grade of tolerance to cold 耐低温级别Seedling mortality (%)赤枯率（%） Grade of tolerance to cold耐低温级别低温条件下磷肥对水稻幼苗生物量的影响由图2可以看出，常温条件下随着施磷量的增加，吉粳81号和长白9号百珠干重呈先升后降的变化趋势，当施磷量为50 g m-2时，两品种百珠干重均达到最大值。而在低温条件下，两品种百珠干重均受到不同程度抑制，且吉粳81号受抑制程度明显小于长白9号，其中与常温未施磷处理相比较，吉粳81号和长白9号分别下降19.12%和27.8%，试着施磷量的增加，尽管其变化趋势与常温相类似，但但磷肥最佳施用量明显高于常温，吉粳81号和长白9号均在施磷量为75 g m-2时干重达到最大值。图2 低温下施用磷肥对水稻生物量的影响Jijing 81 吉粳81号 Changbai 9 长白9号Dry weight of 100 seedlings (g) 白珠干重(g) Normal temperature 常温Low temperature 低温低温条件下磷肥对水稻幼苗SPAD的影响由表3可知，常温条件下，除实磷量25 g m-2以外其他施磷处理两者的SPAD值均高于未施磷处理，且于施磷量50 g m-2时达到最大值，之后继续增施磷肥，两品种幼苗叶片SPAD值明显下降，不同耐寒品种存在明显差异。与常温对照相比较，在低温未施磷肥处理条件下，吉梗81号下降5.97%，而长白9号下降14.5%。随施磷量增加，两者叶片SPAD值下降幅度有逐渐减小趋势，当施磷量为75 g m-2时，吉梗81号和长白9号叶片SPAD下降幅度分别仅为2.9%和4.2%。表3 低温下施磷肥对水稻光合作用的影响Treatments处理方式Jijing 81 吉粳81号Changbai 9 长白9号Normal temperature 常温Low temperature 低温Different small letters indicated significant at 5% level. The same as below.同一列不同小写字母表示5%显著水平差异。下同。低温条件下磷肥对水稻幼苗净光合速率的影响由图3可知，常温下吉粳81号和长白9号净光合速率随着施磷量的增加呈先升后降的变化趋势，且均于施磷量为50 g m-2时达到最大值。而在低温条件下，无论是吉粳81号还是长白9号净光合速率均显著降低，其中吉粳81号净光合速率要高于长白9号，在施磷量为25-75 g m-2时，两者净光合速率随着施磷量增加而上升，并于75 g m-2时达到最大值；当施磷量为100 g m-2时,两品种净光合速率均有所下降，其中长白9号下降幅度明显高于吉粳81号。图3 低温下施磷肥对水稻光合作用的影响Jijing 81 Changbai 9Pn (mol m-2 s-1) 净光合速率（mol m-2 s-1）Normal temperature 常温Low temperature低温低温条件下磷肥对水稻幼苗荧光参数的影响由表4可知，常温下随着施磷量的增加，两品种Fv/Fm略有下降趋势，但各施磷量间差异不显著。低温条件下，与常温相比较，两品种各处理Fv/Fm显著下降，且吉梗81号始终高于长白9号。其中施磷处理后，在施磷量为2575 g m-2，两品种Fv/Fm随施磷量的增加而提高，其中当施磷量为75 g m-2时，吉梗81号和长白9号分别上升11.7%和24.3%。继续增施磷肥，长白9号Fv/Fm显著下降，而吉梗81号虽有下降，但差异不显著。由表4可见，常温条件下不论是否施磷，吉梗81号和长白9号的qP无显著性变化；而在低温条件下，两品种各处理qP均有所下降，但随着施磷量的增加，两品种qP下降幅度减少，两品种均在施磷量为75 g m-2时下降幅达到最小值；继续增施磷肥，两品种qP显著性下降。常温条件下，随着施磷量增加，吉粳81号和长白9号的qN有降低趋势，各处理间变化幅度较小；而低温条件下，不同耐寒品种存在明显差异，与常温比较，吉梗81号的qN明显上升，而长白9号当施磷量在025 g m-2时却有所下降。表4 低温条件下磷肥对水稻幼苗荧光参数的影响Variety 品种Phosphorus application (g m-2) 施磷量 (g m-2) 低温条件下磷肥对水稻幼苗不饱和脂肪酸含量的影响由表5和图4可知，常温条件下，随着施磷量的增加，吉粳81号和长白9号叶片油酸、亚油酸和亚麻酸略有增加，各种处理之间差异不显著，不饱和脂肪酸指数无明显变化。与长白9号相比较，吉粳81号叶片含有的不好和指数要明显高于长白9号。而在低温条件下，与常温相比较，吉粳81号叶片中油酸含量下降，亚油酸和亚麻酸含量升高；长白9号叶片有段和亚油酸含量下降，亚麻酸含量上升。当施磷量在25-75 g m-2时，随着施磷量的增加，两品种叶片油酸含量变化不明显，亚油酸含量降低，不饱和脂肪酸指数明显上升。表5 低温条件下磷肥随水稻不饱和脂肪酸含量的影响Variety 品种Phosphorus Application (g m-2) 施磷量 (g m-2) Normal temperature 常温Low temperature 低温Oleic acid 油酸Linoleic acid 亚油酸图4 低温下施用磷肥对水稻不饱和脂肪酸的影响讨论日本对水稻低温冻害研究较早。我国水稻耐寒性研究始于20世纪7080年代，结合我国各稻区低温冷害特点，制定相应的水稻耐寒性鉴定与评价方法。目前关于水稻苗期耐寒性鉴定主要以赤枯度作为评价指标，本研究通过对低温条件下不同磷肥处理水稻苗期耐寒性进行鉴定，发现不同内冷品种耐寒程度有明显差异，适当增减磷肥可调高水稻幼苗耐寒能力，提升耐寒级别，尤其对非耐寒品种提升作用明显。磷在细胞组成和物质新城代谢过程中发挥着重要作用，并与蛋白质合成、细胞分裂以及细胞生长密切相关。研究表明，植物磷素缺乏会抑制细胞的形成和增殖，从而导致生长和发育停滞（李等人，2007）。而低温同样也会一直植物生长发育，当水稻育苗遇到低温胁迫时，植株鲜重和干重等生物学性状受到明显影响，且不同耐寒品种存在明显差异（李等人，2006）。本研究表明，无论常温还是低温条件下，施磷有助于水稻幼苗百珠干重增加。但在低温下，耐寒品种受抑制程度要小于非耐寒品种，且两品种施磷量最适用量明显提升，这说明适宜的磷用量能在一定程度上增加水稻幼苗素质，提高水稻生物量，缓解低温对育苗的损伤。结果与对西瓜（陈等人，2007）、番茄（李等人，2006）的研究一致。植物光合作用对低温较为敏感。已有研究表明短期低温会导致磷酸化中间产物的积累而细胞质内磷水平降低（赫里等人，1998），抑制叶绿体合成（曾等人，2000）。而叶绿体含量高低与光合作用又有一定关联性（张等人，2000）。本研究结果表明，无论常温还是低温，施磷有助于水稻幼苗叶片叶绿素相对含量及净光合作用率的提高，但施磷过量会引起两者的降低。低温条件下，耐寒品种叶绿素相对含量和净光合速率虽有下降，但始终高于非耐寒品种。常温下最适合磷作用，低温下适当增施磷肥可在一定程度上减缓水稻幼苗叶绿素相对含量的降低程度，维持叶片较高的净光合速率。研究发现Fv/Fm是植物抗冷性敏感指标，在严峻自然条件下，该值则会明显降低（赵等人，200）。在低温胁迫下，冷锻炼的黄瓜幼苗比未经锻炼的幼苗具有较高的非光化学猝灭系数qN（陈等人，1995）。王等人（2005）通过对水稻幼苗的研究发现低温胁迫下耐寒品种可进行较高的光化学猝灭系数qP和非光化学猝灭系数qN过剩光能的耗散，减少反应中心过剩激发能积累，降低对PSII反应中心和天线系统的影响，从而保护植物光合机构使其影响程度降低，而冷敏感品种反应中心和天线系统在低温胁迫下受到严重影响，qP下降快，但qN却随低温时间的延长而下降，导致光合速率下降。而低磷引起植物叶片PSII光能转换和电子传递速率降低，过剩激发能增加（李等人，2004）。本研究结果表明，常温下不同耐寒品种各处理Fv/Fm、qP和qN随施磷量的增加均无明显变化，而在低温下两品种水稻幼苗Fv/Fm、qP、qN均下降，说明低温导致PS II光化学活性受到抑制，使光合机制受到损伤，从而引起水稻幼苗叶片PS II 反应中心转换能力。无论是耐寒品种还是非耐寒品种，低温下适当增施磷肥均可以使水稻幼苗提高光能转化效率，同时通过较高的qP进行荧光猝灭，减轻过剩激发能对反应中心的伤害，减轻热耗散。大量研究表明增加膜脂的不饱和脂肪酸含量可有效降低膜脂的相变温度，且膜脂不饱和脂肪酸成分比例越大，相变温度越低，抗寒能力也就越强（沈等人，1997）。同时抗寒植物一般都具有较高的膜脂不饱和度，可以在较低的温度下保持流动性，保持正常的生理功能。本研究结果表明无论常温还是低温下，耐寒水稻品种具有较高的不饱和脂肪酸指数。因此，其含有较多的不饱和脂肪酸。常温下随着施磷量的增加，不同耐寒性水稻品种不饱和脂肪酸指数均无明显变化，耐寒品种幼苗不饱和脂肪酸指数要明显高于非耐寒品种。而在低温下随着施磷量的增加，适当增施磷肥，无论是耐寒品种还是非耐寒品种不饱和脂肪酸指数均明显上升，说明低温下增施磷肥可通过增加不饱和脂肪酸，增加膜流动性，从而减轻低温对水稻造成的伤害。结论磷在提升低温条件下水稻幼苗耐寒性方面具有非常重要的作用。通过在育苗床土中适当增施磷肥可提升水稻幼苗低温下耐寒级别，增强水稻幼苗素质，减缓水稻幼苗叶绿素相对含量的降低程度，提高净光合速率、光能转化效率以及不饱和脂肪酸指数，从而提高水稻幼苗的耐寒性，且增磷对非耐寒品种的影响要明显高于耐寒品种。目前，吉林东部地区苗期育苗床土平均施磷量在50 g m-2，为防治苗期低温冷害，不仅要注重品种选择，还要改变床土中磷的比例。本研究结果表明，施磷量75 g m-2可有效提高水稻耐寒能力，降低受损程度。Effects of Phosphate Fertilizer on Cold Tolerance and Its Related Physiological Parameters in Rice Under Low Temperature StressHou Li-gang1, 2, Chen Wen-fu1*, Zhao Guo-chen2, Ma Wei2, Qi Chun-yan2, Liu Liang2, and Sun Hong-jiao21 Rice Research Institute, Shenyang Agricultural University, Shenyang 100866, China;2 Rice Research Institute, Jilin Academy of Agricultural Sciences, Gongzhuling 136100, Jilin, ChinaAbstract: The study was designated to explore the physiological mechanism of cold tolerance enhanced by phosphate in rice. An experiment was conducted to investigate the effects of different levels of phosphate fertilizer on cold tolerance and its related physiological parameters in rice seedings (chilling-sensitive cv. Changbai 9 and chilling-tolerant cv. Jijing 81) under low temperature stress. At the same time, the identification of cold tolerance was conducted. Compared with the normal temperature treatment, the relative chlorophyll content, photosynthesis rate, Fv/Fm and qP decreased and index of unsaturated fatty acid increased in rice under low temperature stress. The effect of chilling-sensitive cultivars was more than that of chilling-tolerant cultivars, more phosphorus fertilizer properly improved seedling quality of rice, slowed relative chlorophyll content dropping degree of rice seeding, increased photosynthesis rate, Fv/Fm, qP and index of unsaturated fatty acids, and enhanced the ability to chilling-tolerant cultivars under low temperature. The effect on chilling-tolerant cultivars was significantly higher than that on chilling sensitive cultivars by applying more phosphorus fertilizer. Phosphate regulated photosynthetic physiology and membrane fluidity to reduce injury by low temperature, and increasd the cold tolerance capacity of rice.Key words: low temperature stress, phosphate fertilizer, rice, cold tolerance, physiological parameterIntroductionLocated in the East Asian monsoon region, China is one of the climate vulnerable areas in the world. According to statistics, only in the agricultural field, more than 4 800 million hectares of agricultural production were hit by meteorological disasters each year, which caused serious economic losses in grain production. Cold damage, as a current major agrometeorological disaster, led to Chinas annual losses of up to 50 to 10 billion kg only in rice production (Wang, 2008). Analysis on the meteorological data of the last 50 years (1951 to 2000) in Jilin Province shows that there is an annual average rise in temperature, but in the main rice growing season, regional and stage cold damage phenomena still occur from time to time, and tend to be frequent and severe, and thus result in a year of cold damage (Yu et al., 2003). Phosphorus, one of the necessary nutritional elements for plant growth and development, usually participates in the activities of plant life in multiple ways. The growing number of studies have shown that phosphorus can significantly improve the resistance of the plant, especially in cold tolerance (Wang et al., 1999; Li et al., 2004; Wei et al.,2005; Chen et al., 2007). Years of practice has confirmed that low temperature can affect the rice phosphate absorption and inhibit the growth; while increasing P fertilizer can promote the growth of rice root and help to turn the seedlings green after transplanting, thereby improving rice yield. In recent years, studies on sugar cane (Wei et al., 2003), watermelon (Chen et al., 2007), winter wheat (Li et al., 2004) and tobacco (Wang et al., 1999) plants show that an appropriate increase in P fertilizer plays an important role in improving plant tolerance to cold. However, few studies are conducted on the physiological mechanism of how the increase of P fertilizer can enhance low temperature resistance of rice seedlings. Taking the mechanism as object, this study employed two cold tolerance rice varieties as materials to research the effects of increased P fertilizer on the rice seedlings under low temperature cold tolerance and related physiological characteristics. It aimed to further elucidate the physiological mechanism and provided a theoretical basis for phosphorus fertilizer-centered anti-cold-technical measures.Materials and MethodsTested materialsThe test was conducted in the Breeding Base of Jilin Academy of Agricultural Sciences in 2010, taking different cold tolerance varieties Jijing 81 (chillingtolerance, Jijing 81) and Changbai 9 (chilling-sensitive, Changbai 9) as test materials.Experimental designFirstly, selected the seeds of the same degree in maturity and plumpness and then soaked them for potential bud in mid-April 2010. Secondly, selected seeds with similar bud growth and sowed them evenly in seedling trays which were mixed with different phosphates, with the sowing rate of 200 g m-2 per disk. Five phosphate fertilizer application rates (P2O5)-0, 25, 50, 75 and 100 g m-2 were divided into two groups, one for low temperature processing, another group was under normal temperature control, each group was repeated twice. Finally, irrigated enough water and covered them in the seedling greenhousewith reference to the high yield of rice cultivation practices in Jilin Province.Temperature treatmentWhen the rice seedlings grew up to three leaves, removed the group under low temperature treatment to the artificial climate chamber (LT/ACR-2002, artificial climate chamber control system of Yishengtaihe Company) for continual low temperature stress for 5 days and the average daily temperature was 8; referred to the specific parameter settings in Table 1.The other group stayed outdoor whose temperature data was collected via IRRI weather recorder; the concrete temperature is shown in Fig. 1 (Gongzhuling, 2010). During the processing, the average daily temperature was over 15 and water was given quantitatively.Table 1Temperature and light setting for climate-control chamberFL, Number of fluorescent lamp groups; NL, Number of Na lamp groups.Measurement items and methodsEvaluation of cold tolerance ability of rice seedlingsWith reference to the approaches by Han et al. (2004) and Li et al. (2006) and combined with the actual situation of Jilin Province, the study identified and rated the ability of rice seedlings by evaluating how dry and red of the leaves. The criteria were as the followings: the first gradethe rate of dry and red leaves20%; the second gradethe rate of dry and red leaves was between 21%-30%; the third grade the rate of dry and red leaves was between 31%-40%; the fourth gradethe rate of dry and red leaves was between 41%-50%; the fifth gradethe rate of dry and red leaves was between 51%-60%; the sixth grade the rate of dry and red leaves was between 61%-70%; the seventh gradethe rate of dry and red leaves was between 71%-80%; the eighth gradethe rate of dry and red leaves was between 81%-90%; and the ninth gradethe rate of dry and red leaves was90%.Determination of biomassAfter the processing, selected 100 seedlings from each group respectively, rinsed them with distilled water, fixed them at 105 for 15 min and then dried them to get constant dry weight of 100 seedlings (g).Determination of the SPADThe amount of chlorophyll content was measured by chlorophyll meter (SPAD-502 type). Gripped the center of the uppermost fully expanded leaves with chlorophyll meter to evaluate the SPAD value. Took 10 seedlings for one group and then got their average SPAD value. Repeated the process for three times.Determination of net photosynthetic rateUsed LI-6400 portable photosynthesis system manufactured by American LI Company to determine the net photosynthetic rate of rice leaves (Pn).Determination of chlorophyll fluorescence parametersThe chlorophyll fluorescence parameters of rice leaves were determined by Germany WALZ PAM 2500 portable pulse modulated chlorophyll fluorescence analyzer. Measurement methods referred to Genty et al. (1989). Took uppermost fully expanded leaves, made dark adaptation before the measurement for 20 min, and then opened the measuring light and got the original fluorescence of F0, then opened the saturation pulse of light to get the maximum fluorescence Fm and maximal photochemical efficiency of Fv/Fm. Then opened the photochemistry, made light adaptation for 20 min and then got the fluorescence parameters of the measured light existence of the maximum fluorescence Fm, among which, Fv/Fm, qP, and qN were derived by the following formula:Fv/Fm (maximum photochemical efficiency)= (Fm-F0)/Fm;qP (variable photochemical devoid)=(Fm-Ft)/ (Fm-F0);qN was (non-photochemical fluorescence quenching coefficient)=Fm/Fm1.Determination of unsaturated fatty acids in leaves The oleic acid, linoleic acid and linolenic acid content were measured with reference to the methods by Yu et al. (1996). The formula was that the index of unsaturated fatty acids (UFA)=oleic acid content (mol%)1+linoleic acid content (mol%)2+linolenic acid content (mol%)3.ResultsEffects of phosphate on cold tolerance ability of rice seedlings at low temperatureThere was an obvious difference in cold resistance ability among different rice varieties (Table 2). Under low temperature conditions, with no application of phosphate, the rate of dry and red leaves of Jijing 81 was 17.6% and its resistance ability was the first grade. In contrast, the rate of dry and red leaves of Changbai 9 was 31.2% and its resistance ability was the thirdgrade. After adding phosphate to 0-75 g m-2, the rate of dry and red leaves of both varieties declined but the rate of Jijing 81 reduced a little, Changbai 9 changed significantly. As for their cold tolerance abilities, Jijing 81 had no change and Changbai 9 increased upto grade 1. When phosphate was added to 100 g m-2, the rate of dry and red leaves of both Jijing 81 and Changbai 9 increased, between which there was an obvious rise in Changbai 9 and its cold tolerance ability declined from grade 1 to grade 2.Table 2Cold tolerance identification of rice at seedling stage under low temperature stressEffects of phosphate on rice seedling biomass under low temperature conditionIt could be seen from Fig. 2 that under normal temperature, the dry weight of 100 seedlings tended to increase first and then decreased with the increase of phosphate amount. When the phosphate amount reached 50 g m-2, both varieties reached their maximum in dry weight.While under low temperature condition, both dry weights were subject to vary