拟南芥硝酸调控基因NRG2.10的功能研究

Nitrate is an essential nutrient, and is involved in many adaptive responses of plants, but how such nitrate-specific mechanisms are regulated at the molecular level is poorly understood. This research used the T-DNA insertion mutants of NRG2.10, which is the homology gene of a new nitrate regulatory gene NRG2, to investigate the function of NRG2 in nitrate regulation. This project is mainly to study the function of NRG2.10 on nitrate regulation, nitrate translation and nitrate assimilation aspects. First we tested the nitrate induction of nitrate inducible genes in mutants after nitrate treatment to determine if NRG2.10 is a nitrate regulatory gene, and its affects in molecular levels. Second we measured the nitrate contents in mutants to work out if NRG2.10 will affect the nitrate accumulation in mutants. Then we tested the amino acid content in mutants to investigate if NRG2.10 affected the nitrate assimilation in mutants. We also tested the expression pattern of NRG2.10 in Arabidopsis use QPCR and GUS staining, and the subcellular localization of NRG2.10 use GFP fusion protein. And we will make double and triple mutants of NRG2.10 and NRT1.1 and NLP7 genes to study the relationship of NRG2.10 and other known nitrate regulatory genes. We will also use yeast two hybrid and BiFC to work out if the NRG2.10 will interact with the other nitrate regulate genes. This research will establish the key role of NRG2.10 in central nitrate signaling and offer insights into the mechanism of nitrate regulation in plants. In addition, our project provides the important insights into the functions of NRG2 gene family in Arabidopsis.

氮素对植物生长发育具有重要的作用，而硝酸是植物吸收氮素的主要形式，但目前对于硝酸调控机理以及硝酸信号分子调控网络的研究还很少。本项目将研究一个新的硝酸调控基因NRG2.10，阐明它在硝酸调控方面所起的重要作用。首先通过检测硝酸诱导的基因和调控基因的表达来分析NRG2.10在分子水平对硝酸调控的影响；其次通过检测硝酸含量等来研究NRG2.10对硝酸吸收利用的影响；再次检测NRG2.10的组织表达和亚细胞定位来研究基因的表达和定位。本研究还将通过构建NRG2.10与硝酸调控基因NRT1.1和NLP7的双突变体和三突变体等，研究NRG2.10与NRT1.1和NLP7之间的关系，并且验证NRG2.10与已知硝酸调控基因之间是否存在互作。本研究可以进一步健全硝酸调控基因网络，为研究NRG2基因家族提供了理论基础，为阐明硝酸调控的分子机理提供重要依据。

氮素对植物生长发育具有至关重要的作用，而硝酸是多数植物对氮素吸收利用的主要形式，因而研究植物对硝酸的吸收利用规律，进而丰富健全硝酸调控网络，对于提高作物的氮素利用率、减少环境污染具有重要意义。本研究找到一个新的硝酸调控基因NRG2.10，在硝酸诱导后，发现其突变体对硝酸的响应程度明显比野生型弱，在突变体中硝酸诱导基因的诱导量显著低于野生型，表明此基因是一个新的硝酸调控基因。进一步研究发现此NRG2.10基因突变体根中的硝态氮含量较野生型低，地上部硝态氮含量高于野生型，而造成上述现象的主要原因是由于硝酸转运基因NRT1.8表达上调和硝酸同化基因GLN1.1表达上调。根据QPCR检测结果，发现NRG2.10基因主要在根中和叶中表达，但是叶中表达量低于根中，亚细胞定位显示此基因编码的蛋白主要在细胞核中表达。对nrg2.10/ nrg2.11双突变体的研究发现，双突变体植株长势小于野生型且根短于野生型，以上结果均证明NRG2.10基因有重要的硝酸调控功能，且NRG2家族对植物的生长有重要作用，我们的研究对于认识这个基因的功能，建立和丰富硝态氮调控的分子网络，进而加深对硝酸调控基因网络的研究具有重要意义。