氮效率遗传改良是提高氮肥利用效率的重要途径，而茎叶氮素向籽粒的高效转运是玉米高效利用氮素的重要生理机制之一。前期研究结果表明，先玉335杂交种具有氮转移效率高的特性，其亲本在叶片保绿度、氮转移效率、籽粒蛋白质浓度等性状存在显著差异。利用先玉335双亲组配DH分离群体，对上述性状定位到一个主效QTL-qNT5.04。结合精细定位与亲本转录组差异分析，确定了候选基因，该基因编码一个Clp家族蛋白酶。本项目拟利用候选基因关联挖掘有利等位基因变异，利用Crispr/Cas9系统与超表达对候选基因进行功能验证。通过生理指标，亚细胞定位、组织特异性表达、酵母双杂等生理与分子生物学手段解析该基因的生理生化功能，建立玉米叶片氮转移与产量形成、籽粒蛋白质积累的分子调控网络。最终，阐明该基因的进化关系，为协同提高玉米产量、氮效率及籽粒蛋白含量提供基因资源。

Genetic improvement of nitrogen efficiency is an important way to improve nutrient efficiency, and high nitrogen remobilization efficiency of stems and leaves is an important physiological mechanism of nitrogen efficiency in maize. Previously, our results showed that Xianyu 335 showed high efficiency in nitrogen remobilization from stem and leaf to grain, reducing nitrogen residue, and achieving high yield, high nitrogen efficiency and high grain protein simultaneously. We also found that there were significant differences in leaf greenness, nitrogen remobilization efficiency and grain protein concentration between the two parents of Xianyu335. One major QTL-qNT5.04 was identified using DH segregation populations formed by selfing Xianyu335. The QTL was cloned by combining QTL fine-mapping and RNA-seq analysis of the parents of Xianyu335 and the candidate genes encode a Clp family protease. Based on these completed work, this project intends to detect favorable allele variance which was mined by candidate gene association study and the function of candidate gene was verified by Crispr/Cas9 system and over-expression. Physiological and molecular biological methods such as subcellular localization, tissue specific expression, yeast heterozygosity were used to analyze the physiological and biochemical functions of the gene, and a molecular regulatory network was established for nitrogen remobilization and yield formation in maize leaves and protein accumulation in grains. Finally, the evolutionary relationship of the gene was elucidated to provide genetic resources for synergistic improvement of maize yield, nitrogen efficiency and grain protein content.