减量施氮下扩张蛋白基因调控冬小麦形态和生理可塑性的机理

The problem of excessive nitrogen application is common in winter wheat production in China. Excessive nitrogen application not only did not improve wheat yield, but also caused waste of resources and environmental pollution. At present, it has been clear that crop morphological and physiological plasticity is an important factor in reducing yield loss and improving resource utilization efficiency in stress, and crop plasticity is closely related to the expression and regulation of expansin gene family. However, the mechanism of expansin gene family's expression regulation on plasticity and nitrogen uptake and utilization in winter wheat organs under low nitrogen level is still unclear. The potential effect of EXPB23 gene expression on the plasticity of Winter Wheat under nitrogen gradient has been preliminarily explored by project applicants. This project intends to carry out nitrogen gradient test under field and indoor conditions to investigate the morphological and physiological parameters of winter wheat, the law of nitrogen uptake and utilization, the expression of different expansive protein genes in different organs and the content of expansive protein. gene silencing technology is used to verify the function of differentially expressed genes, and the expansive protein is identified by making full use of the prediction and annotation function of bioinformatics software. The mechanism of regulating photosynthetic physiology, organ growth and development, and nitrogen uptake and utilization. The results provide theoretical guidance for rational reduction of nitrogen application, making full use of morphological and physiological plasticity of winter wheat, and improving nitrogen use efficiency.

我国冬小麦生产中普遍存在施氮量过大的问题。过量施氮不仅没有提高小麦产量，而且造成了资源浪费和环境污染。目前已明确作物的形态和生理可塑性是逆境中作物减少产量损失、提高资源利用效率的重要因素，且作物可塑性与扩张蛋白基因家族的表达及调控有密切关系。但是低氮下扩张蛋白基因家族成员在冬小麦各器官中表达调控对可塑性及氮素吸收利用的影响机理尚不明确。项目申请人已初步探索了氮梯度下EXPB23基因表达量对冬小麦可塑性的潜在影响。本项目拟在大田和室内条件下开展施氮梯度试验，调查冬小麦形态生理指标、氮素吸收利用规律、各器官中不同扩张蛋白基因的表达量及扩张蛋白含量，使用基因沉默技术进行差异表达基因的功能验证，并充分利用生物信息学软件的预测和注释功能，探明扩张蛋白调控光合生理、器官生长发育及氮素吸收与利用的机理。研究结果为合理减量施氮，充分利用冬小麦形态和生理的可塑性，提高氮肥利用效率提供理论指导。

背景：小麦产量与氮肥施用量密切相关。在不影响小麦产量的情况下减少氮肥的施用是一项具有挑战性的任务。探明小麦与氮亏缺有关的代谢、生理和形态响应，以及相关信号通路和基因，对通过农艺或育种手段提高氮肥利用效率至关重要。.主要研究内容：开展了施氮梯度试验，调查冬小麦形态生理指标、各器官中不同Expansin基因及其它相关基因的表达量及蛋白含量，充分利用生物信息学软件的预测和注释功能，探明Expansin及其它相关基因调控光合生理、器官生长发育及氮素吸收与利用的机理。使用CRISPR/Cas9技术获得Expansin候选基因TraesCS1A02G212400的F2代过表达突变体，并做氮亏缺下的表型鉴定。.重要结果：氮亏缺抑制了小麦叶片的生长，促进了小麦根系的生长。氮亏缺下小麦叶片的光合速率和气孔导度先增加后降低。根系中24个Expansin（EXPB3、31EXPA31、EXPA14、EXPA24和EXPA9等）基因表达量的提高可能与小麦根长、根体积、根表面积的增加有关。另外发现，根系中4个NRT基因表达量的提高可能促进小麦根系氮素的转运；叶片中2个NAR和9个WRKY基因表达量的下降可能与株高、叶面积的降低有关；叶片中11个PSB基因表达量的提高可能促进小麦光合作用。Expansin、WRKY、NAR、NRT和PSB差异基因可分为3，3，2，3，12个亚族，分别分布在小麦的17，16，3，4，13条染色体上。根中表达量显著提高的24个Expansin基因富集在与细胞壁延展功能相关的组分中。使用CRISPR/Cas9技术获得了Expansin候选基因TraesCS1A02G212400的过表达突变体F2代植株。.关键数据及其科学意义：Expansin（EXPB3、31EXPA31、EXPA14、EXPA24和EXPA9等）、NRT2（NRT2.1、NRT2.2和NRT2.3）、PSBR基因是本研究提供的与氮高效吸收利用相关的参考基因。CRISPR/Cas9过表达EXPB3（TraesCS1A02G212400）基因的F2代突变体经过初步鉴定能够提高小麦形态和生理可塑性，从而提升植株对氮亏缺的适应能力，可作为今后在基因、表型可塑性、资源利用效率关系的相关研究中重要参考基因。