干湿交替驱动水稻根表铁膜形成的生理与分子机制

Iron plaque on root surface is an important physical barrier and nutrient pool. Soil drying and wetting alternation is a traditional management strategy in rice production. Studying the formation mechanism of iron plaque can provide scientific basis for improvement of rice quality and production. In this study rice varieties differing in iron plaque quantity will be used as experimental materials. Equipment and methods including DCB method, electron microscope, X-ray photoelectron spectrometry etc. will be used to characterize the type, quantity and distribution of iron plaque. Microelectrode and chromatograph mass spectrometer will be used to measure rhizospheric Eh value and dynamics of oxidative substances. The above results will contribute to elucidating rhizospheric characteristics driven by wetting and drying alteration. Then histochemical staining and colorimetric method will be used to determine the activities of antioxidant enzymes, reactive oxygen species etc. so as to understand the physiological mechanism of iron plaque formation driven by drying and wetting alteration. Based on the above results, high throughput sequencing technique and pathway analysis will be used to isolate differential expressed genes. Transgenic plants will be made by using agrobacterium tumefaciens mediated transformation. The function of a candidate gene in forming iron plaque will be evaluated during wetting and drying alteration. Thus the molecular mechanism of iron plaque formation will be explored. The above results will play an important role in elucidating formation of iron plaque and provide scientific evidences for breeding new rice varieties.

根表铁膜是水稻适应水生环境的重要物理屏障和养分库。稻田干湿交替过程是水稻生长过程中常见的管理方法，研究稻田干湿交替驱动水稻根表铁膜的形成机理，对阻止有害重金属和有机污染物通过水稻根系进入籽粒具有重要意义。本项目拟利用铁膜数量差异显著的水稻品种为材料，利用DCB方法、电镜、X射线与能谱分析仪等表征水稻根表铁膜的种类、数量与分布；利用微电极、色谱与质谱联用仪等测定水稻根际Eh值和氧化性物质的动态变化，明确干湿交替驱动根表铁膜形成的根际特征。然后，借助组织化学染色和比色法等测定水稻根系抗氧化酶活性、活性氧累积等，明确干湿交替驱动根表铁膜形成的生理机制。在此基础上，利用高通量测序和Pathway显著性富集分析，分离干湿交替诱导铁膜形成的差异表达基因；借助农杆菌介导转化，获得转基因材料；并评价转基因材料在铁膜形成过程中的作用。该研究结果对阐明水稻根表铁膜的形成机制和水稻新品种选育具有重要科学意义

根表铁膜是水稻适应水生环境、抵抗逆境胁迫的重要屏障。干湿交替能明显诱导水稻根表铁膜的形成，然而并不清楚其诱导机理。本项目研究了干湿交替诱导水稻根表铁膜形成的分子生理机制，发现：干湿交替明显影响水稻根际氧化还原电位（Eh）。落干时，根际Eh值上升，淹水时下降。淹水处理时，根表无定形铁含量升高，结晶铁下降。干湿交替增加根际铁膜结晶度。适度干湿交替次数（分蘖期2次、孕穗期1次、成熟期1次）明显增加水稻根系氧化性物质（过氧化氢）累积，提高根系氧化力，促进铁膜在根表累积。与淹水相比，干湿交替时，水稻根表铁膜中的赤铁矿、磁铁矿、针铁矿、纤铁矿较多，而水铁矿较少。.水稻根系基因差异表达谱分析发现，3822个差异表达基因参与了干湿交替和铁膜形成过程。102个差异表达基因参与了氧化还原反应过程，30个基因上调表达，72个基因下调表达。干湿交替处理增加了水稻根系乙醇酸氧化酶基因和过氧化物酶基因的表达，与对照相比，干湿交替处理下，水稻根系OsGLO4基因和OsPrx04基因的表达量分别是对照的2.7倍和1.9倍。组织特异性分析发现，OsGLO4在水稻根和茎中表达丰度较高。OsPrx04在水稻根、茎、叶三部位均有表达。.转基因材料分析发现，干湿交替条件下，OsGLO4超表达材料（OsGLO4-OX）根表铁膜数量是其野生型材料135-167%，而OsGLO4缺失突变体(OsGLO4-KO)根表铁膜数量是其野生型材料61.5-75.6%。对于OsPrxO4基因来说，干湿交替处理的OsPrxO4-OX根表铁膜数量明显大于长期淹水处理。OsPrxO4-KO的铁膜数量没有明显变化，但结晶态铁含量上升。OsPrx04-OX和OsPrxO4-KO材料在干湿交替处理后的结晶度大于野生型，但在长期淹水时没有明显差异。干湿交替时，水稻根系OsGLO4基因和OsPrx04基因通过增加过氧化氢累积，提高根系氧化力，从而诱导更多铁膜累积。这些研究结果对指导水稻生产具有重要现实意义。.受该项目资助，本课题组成员发表了中文学术论文7篇，英文SCI收录论文4篇，其中1区SCI论文2篇。同时，参加国内和国际学术会议各1次，发表会议论文2篇。申报国家发明专利2项，授权1项。培养博士研究生2名，硕士研究生2名。