水稻根特异表达基因Os03g01700的启动子研究和应用

Most studies on organ growth control investigate growth of aerial organs, whereas roots are only rarely considered. However, Plant roots perform many essential functions, including water and nutrient uptake, storage, and transportation. The root system architecture determine the plant's ability to access water and nutrients, factors that limit growth and, thus, yield in plants. Root-specific promoters and their cis-elements are important to drive root-specific gene expression and are used to improve nutrient and water uptake, root growth in plants. Many genes described as "Intrinsic Yield Genes"(IYGs) in plants, when mutated or ectopically, cause formation of larger structures, such as leaves or roots and, hence, more biomass. More recent reports state that IYGs, which is regulated by the root-specific promoter, promote a large root system, improve drought tolerance, nutrient efficiency of crop plants, whereas the growth and development of the shoots have not been negatively affected. Rice is one of the most important food crops in world. Rice productivity is limited by low Pi availability in cultivated areas worldwide. To improve root growth, thus, rice yield under Pi-deficient conditions, it is essential to understand how the molecular machinery that determines yield parameters operates. Several Pi-responsive transcriptional factors enhance root growth when ectopically (over)expressed. There was a significant increase in primary and adventitious root lengths of the OsMYB2P-1-overexpressing plants controlled by CaMV 35S promoter under Pi-deficient conditions compared with wild-type plants, whereas the reduced expression of OsMYB2P-1 led to suppression of the growth rate of primary root and adventitious roots. Therefore, this work demonstrates that manipulation of OsMYB2P-1 might be a feasible approach to genetically engineer plants with enhanced root system. However, Overexpression of OsMYB2P-1 controlled by the constitutive promoter leads to the overaccumulation of Pi in shoots under Pi-sufficient conditions. Therefore, the detrimental consequences of the overaccumulation of Pi in shoots under Pi-sufficient conditions need to be avoided. Here, the root-specific elements in the promoter of root-specific expression gene Os03g01700 and a DNA-binding protein binding to the root-specific element will be identified. Meanwhile, overexpression of OsMYB2P-1 under control of the promoter of root-specific expression gene Os03g01700 will be employed to understand how root-specific expression of OsMYB2P-1 allows the production of transgenic plants with an enlarged root system without negative side effects in the aerial organs. It will also show that plants with an increased root system have an improved Pi uptake, increased biomass production under nutrient limitation. This will establish an approach for modulating root system architecture and might be useful for generating crop plants optimized to grow in difficult agricultural environments.

植物器官生长的研究主要集中在地上部，根中很少涉及。根系构型决定了植物吸收水分和营养的能力，从而制约着产量的提高。根特异启动子及其特异元件在研究植物水分和养分的吸收、根系生长等方面具有重要意义。应用根特异启动子调控植物器官生长相关基因在根系生长，提高水分和养分等方面取了显著成效。 水稻是重要的粮食作物，磷缺乏已成提高水稻产量的限制因素。OsMYB2P-1是水稻MYB2磷响应基因１，组成型超表达OsMYB2P-1的转基因材料根系发达，根生物量增加；但在高磷条件下，OsMYB2P-1超表达材料地上部表现磷中毒。为了改良水稻根系生长，提高水稻根系对磷的吸收和利用效率，避免使用组成型启动子带来的不利影响。本项目将对水稻根特异表达基因Os03g01700启动子的特异元件及结合蛋白进行研究，同时，利用其启动子调控OsMYB2P-1在水稻根中特异表达，实现根构型改良，提高磷的利用效率。

根特异启动子及其特异调控元件在研究植物水分和养分的吸收、根系生长及构型等方面具有重要意义。研究显示，应用根特异启动子调控植物相关基因在根系中表达，促进植物的根系生长，根构型改良，实现理想株型，提高水分和养分等方面取了显著成效。..水稻是重要的粮食作物，磷缺乏已成提高水稻产量的限制因素。OsMYB2P-1是水稻MYB2磷响应基因１；但在高磷条件下，OsMYB2P-1超表达材料地上部表现磷中毒。为了改良水稻根系生长，提高水稻根系对磷的吸收和利用效率，避免使用组成型启动子带来的不利影响。本研究对水稻根特异表达基因Os03g01700启动子的特异元件进行了分析，同时，利用水稻根特异表达基因Os03g01700的启动子调控OsMYB2P-1在水稻根中特异表达。..项目对水稻根特异表达基因Os03g01700的启动子启动子-2013bp～-1475bp和-1077bp～-561bp区特异区段进行缺失分析，在-1077bp～-561bp区段，根据实验结果，无法确认具体的特异区段；在-2013bp～-1475bp区段，大约148bp的长度上，可能存在根特异的调控元件；对此区段再作进一步的缺失分析，设计了3P-1939bp::GUS，进一步缩小了特异区段的位置，确定其位于-1939bp～-1865bp，74bp的长度范围内。后续对其进行了突变分析，在构建的3PM1----3PM7材料中，3PM4属于根特异表达元件，突变后GUS活性降低；3PM6属于叶特异表达的调控元件；突变后GUS活性上调；3PM3在在所有突变材料中，GUS活性最低。结合功能获得材料的数据，可以确定特异元件。..获得ＰOs03g01700:OsMYB2P-1、ＰOsDX1:OsMYB2P-1 、35S:OsMYB2P-1阳性超表达转基因材料。在磷生理实验中，进行了高磷及低磷生理实验分析。在高磷状态下，3P::OsMYB2P-1与野生型很接近，地上部分有轻微的磷中毒现象，说明根的特异超量表达使其获得具有更强的磷吸收利用能力；在低磷条件下，磷吸收能力比野生型稍强。其在各项表型参数，如株高、分蘖及根长方面并没有优于野生型。项目进一步分析了POs03g01700:OsMYB2P-1转基因材料中磷响应基因OsPHR2、OsPT2、OsIPS1的表达情况，为研究水稻根系和磷信号途径提供研究材料。