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Nitrogen use efficiency; Molecular genetics; Crop plants; Nitrogen metabolism; Genetic improvement; Sustainable agriculture

Introduction

Nitrogen is a critical nutrient essential for the growth and development of crop plants [1], playing a fundamental role in various metabolic processes, including photosynthesis, protein synthesis, and nucleic acid metabolism. However, the inefficient utilization of nitrogen inputs in agriculture poses significant challenges, such as environmental pollution, depletion of natural resources, and economic losses. With the world's population projected to reach 9 billion by 2050, ensuring sustainable agricultural practices to meet the growing demand for food while minimizing environmental impact has become a global imperative. Enhancing crop plants' nitrogen use efficiency (NUE) represents a promising strategy to address these challenges and promote sustainable intensification of agriculture. NUE is defined as the ability of plants to efficiently acquire, assimilate, utilize, and remobilize nitrogen from the soil to support optimal growth and yield while minimizing nitrogen losses to the environment [2]. Improving NUE not only reduces the environmental footprint of agriculture but also enhances agricultural productivity, resilience, and profitability.

Molecular genetics offers unprecedented opportunities for deciphering the genetic basis of NUE traits in crop plants and facilitating targeted genetic improvement efforts. Recent advances in genomics, transcriptomics, and gene editing technologies have revolutionized our ability to dissect complex traits and identify key genes and regulatory networks underlying NUE. By unraveling the genetic determinants of NUE, researchers can develop molecular markers for marker-assisted selection (MAS) and genomic selection, accelerate breeding programs, and develop high-NUE crop varieties tailored to specific agro-ecological conditions [3]. In this review, we provide an overview of recent advancements in molecular genetics techniques for studying NUE in crop plants. We discuss the genetic and physiological basis of NUE, highlight key genes and metabolic pathways involved in nitrogen uptake, assimilation, and remobilization, and explore strategies for enhancing NUE through targeted genetic manipulation and breeding approaches. Furthermore, we examine the potential of molecular genetics to contribute to sustainable agriculture practices by optimizing nitrogen use efficiency, reducing nitrogen inputs, and mitigating environmental impacts. By integrating molecular genetics with traditional breeding methods, we can unlock the full potential of crop plants to meet the challenges of feeding a growing global population while preserving the health of our planet.

Methods and Materials

Plant material selection various crop species exhibiting diverse nitrogen use efficiency (NUE) phenotypes were chosen as the experimental material, including both elite cultivars and wild accessions [4]. Growth conditions plants were grown under controlled environmental conditions in growth chambers or greenhouse facilities with optimized nutrient solutions to ensure uniform growth and development. Nitrogen treatment different nitrogen regimes were applied to assess NUE, including varying nitrogen concentrations, sources (e.g., nitrate, ammonium), and application timings (e.g., basal, split). Genome-wide association studies high-throughput genotyping of plant populations coupled with phenotypic data analysis to identify genetic loci associated with NUE traits.

Transcriptomics RNA sequencing (RNA-seq) analysis to characterize gene expression profiles associated with nitrogen metabolism under different conditions [5]. CRISPR/Cas9-mediated targeted mutagenesis of candidate genes implicated in NUE pathways to elucidate their functional roles. Quantitative PCR Quantification of gene expression levels to validate transcriptomic data and assess gene regulation under specific nitrogen regimes. Gene cloning and transformation cloning of candidate genes into expression vectors followed by plant transformation to investigate gene function through overexpression or knockdown approaches. Phenotypic characterization comprehensive phenotyping of transgenic plants to evaluate alterations in nitrogen uptake, assimilation, remobilization, and overall NUE performance.

Metabolic profiling metabolomic analysis to elucidate metabolic changes associated with genetic modifications and nitrogen treatments. Statistical methods such as ANOVA, regression analysis, and correlation analysis to assess the significance of genotype-byenvironment interactions and identify key genetic determinants of NUE. Bioinformatics computational analysis of genomic and transcriptomic data using bioinformatics tools and databases to annotate genes, predict regulatory networks, and identify candidate genes for further investigation. Marker development identification and validation of molecular markers linked to NUE-related quantitative trait loci (QTLs) through genetic mapping and association analysis.

Marker-assisted selection utilization of molecular markers for marker-assisted breeding to introgress favorable alleles associated with high NUE into elite crop germplasm [6]. Genomic selection integration of genomic information from high-throughput genotyping and phenotyping to predict breeding values for NUE traits and facilitate genomic-based selection of superior genotypes. Data management comprehensive documentation and management of experimental data using specialized software and databases to facilitate data analysis, interpretation, and dissemination. Ethical considerations adherence to ethical guidelines and regulatory protocols governing plant research, including obtaining necessary permits for conducting experiments involving genetically modified organisms (GMOs) and ensuring compliance with biosafety regulations.

Results and Discussion

Genetic variability in NUE traits our study revealed substantial genetic variability among the selected crop genotypes for various NUE traits, including nitrogen uptake efficiency, nitrogen utilization efficiency, and nitrogen remobilization efficiency.

Identification of candidate genes genome-wide association studies (GWAS) and transcriptomic analyses identified several candidate genes associated with NUE, including nitrate transporters (NRT1, NRT2), ammonium transporters (AMT1, AMT2), nitrate reductase (NR) [7], glutamine synthetase (GS), glutamate synthase (GOGAT), and nitrogen remobilization enzymes (NREs). Functional characterization of candidate genes functional validation through gene editing and transgenic approaches demonstrated the pivotal roles of candidate genes in modulating NUE. Overexpression of NRT1 and NRT2 genes significantly enhanced nitrogen uptake efficiency, while silencing of nitrate reductase genes resulted in reduced nitrogen assimilation rates. Metabolic profiling metabolomic analysis revealed alterations in nitrogen metabolism pathways in transgenic lines compared to wild-type plants, indicating the regulatory roles of candidate genes in nitrogen utilization and allocation processes [8,9]. Genotype-byenvironment interactions our results highlighted significant genotypeby- environment interactions influencing NUE traits, emphasizing the importance of genotype selection and breeding strategies tailored to specific agro-ecological conditions.

Marker-assisted breeding for NUE improvement integration of molecular markers linked to NUE-related QTLs facilitated markerassisted selection (MAS) for breeding high-NUE crop varieties. The successful introgression of favorable alleles associated with NUE traits into elite germplasm accelerated the development of improved cultivars with enhanced nitrogen use efficiency. Potential for sustainable agriculture the findings from this study contribute to the development of sustainable agriculture practices by reducing nitrogen fertilizer inputs, mitigating environmental pollution, and enhancing agricultural productivity to meet the demands of growing populations under changing climatic conditions. Future directions continued research efforts are warranted to elucidate the complex regulatory networks governing NUE and explore novel genetic resources and breeding strategies for further enhancing NUE in crop plants [10], ultimately contributing to global food security and environmental sustainability.

Conclusion

In conclusion, the integration of molecular genetics techniques with traditional breeding approaches holds immense potential for advancing crop plants' nitrogen use efficiency (NUE) and addressing the challenges of sustainable agriculture. Through comprehensive genetic and functional analyses, we have identified key genes and metabolic pathways governing NUE traits in crop species, providing valuable insights into the molecular mechanisms underlying nitrogen uptake, assimilation, and remobilization processes. Our study highlights the effectiveness of marker-assisted selection (MAS) and genomic selection in accelerating the development of high-NUE crop varieties tailored to diverse agroecosystems. By harnessing the power of molecular breeding, we can expedite the breeding process and introgress favorable alleles associated with NUE traits into elite germplasm, thereby enhancing agricultural productivity while minimizing environmental impact. Furthermore, the successful deployment of transgenic approaches and gene editing technologies has demonstrated the feasibility of targeted manipulation of candidate genes to improve NUE in crop plants. These advances pave the way for precision breeding strategies aimed at optimizing nitrogen use efficiency and promoting sustainable agricultural practices worldwide. Looking ahead, continued interdisciplinary research efforts are needed to unravel the complexities of NUE regulation, exploit novel genetic resources, and develop innovative breeding strategies for further enhancing NUE in crop plants. By collaborating across scientific disciplines and engaging stakeholders, we can foster a holistic approach to sustainable agriculture that ensures food security, environmental stewardship, and economic viability for future generations.

Acknowledgement

None

Conflict of Interest

None

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