| Nitrogen is one of the essential elements for plant growth and development.It is an important component of proteins,enzymes,and chlorophyll,as well as crop yield.In agricultural production,nitrogen fertilizer is mainly used to ensure the nitrogen demand of crops,and the excessive use of nitrogen fertilizer also brings about environmental eutrophication.Therefore,the rational application of nitrogen fertilizer and the improvement of nitrogen utilization efficiency are of great significance for the sustainable development of agriculture in China.The Cultivated rye(Secale cereal L.)is an important food and feed crop in the Gramineae family,belonging to the rye genus.It has characteristics such as low nitrogen tolerance,high tillering ability,fast growth speed,strong stress resistance,and strong adaptability.The NRT1.1 genes have been proven to be root nitrate sensors in Arabidopsis and rice,responsible for absorbing and transporting nitrate.However,research works on the NRT1.1 gene related to rye have not been reported.Exploring the function of the NRT1.1gene in rye is of great significance for improving nitrogen absorption and transport in rye and improving nitrogen utilization efficiency.Therefore,this study used Weining rye sequenced in our laboratory as the material to explore the effect of nitrogen stress on its growth.The main results of exploring the function of ScNRT1.1 through q RT-PCR,subcellular localization,and transgenic technology are as follows:Nitrogen stress significantly inhibited the aerial growth of rye,resulting in smaller leaves,higher yellowing,and reduced aerial fresh weight.However,nitrogen stress stimulated root growth within a certain range,resulting in a significant increase in root to shoot ratio and root length.Under nitrogen stress,chlorophyll significantly decreased,while nitrate and total nitrogen contents in both the aerial and root parts significantly decreased.Transcriptome analysis showed that nitrogen stress significantly affected the absorption and metabolism of nitrogen in rye roots.There are a total of 5 NRT1.1 genes in rye roots that respond to nitrogen stress.In this experiment,2 NRT1.1 genes with increased expression levels(ScNRT1.1-1 and ScNRT1.1-3)were selected for exploration.Both ScNRT1.1-1 and ScNRT1.1-3 contain the conserved domain PTR2 of the NPF family protein.ScNRT1.1-1 protein has higher homology with rice Os NRT1.1a protein,while ScNRT1.1-3 has higher homology with wheat Ta NRT1.1 protein.It is speculated that they may have similar functions.Through q RT-PCR detection,it was found that ScNRT1.1-1and ScNRT1.1-3 were both highly expressed in the roots.During the whole process of nitrogen stress,the expression of ScNRT1.1-1 in rye root increased significantly,while the expression of ScNRT1.1-3 increased significantly at multiple time points of nitrogen stress.In conclusion,rye responds to nitrogen stress by inducing the expression of different genes at different time points,and ScNRT1.1-1 may play an important role in this process.Construction of subcellular localization recombinant vectors and transient expression in tobacco showed that ScNRT1.1-1 and ScNRT1.1-3 were both localized on the cytoplasmic membrane,suggesting that they may have the functional ability to transport ions.In the ectopic expression of ScNRT1.1-1 into Arabidopsis Col-0 and At NRT1.1 mutants(chl1-5),the root length,fresh weight,chlorophyll content,nitrate nitrogen content,and NR activity of ScNRT1.1-1 transgenic lines were significantly increased under low nitrogen conditions compared to the wild-type Col-0.Therefore,it is preliminarily inferred that ScNRT1.1-1 can promote the absorption and transport of nitrate in Arabidopsis.This study analyzes the expression patterns of two NRT1.1 genes in rye and explores the function of ScNRT1.1-1,providing reference for studying the mechanism of nitrate absorption and transport in rye,and also providing theoretical basis for genetic improvement of efficient nitrogen utilization in rye. |
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