Ammonium Transport Research Articles

Ammonium Transporter 1 (AMT1) Gene Family in Pomegranate: Genome-Wide Analysis and Expression Profiles in Response to Salt Stress

Understanding the ammonium (NH4+) uptake and transport systems, particularly AMT1 genes, is important for plant growth and defense. However, there is a lack of research on identifying and analyzing AMT1 genes in pomegranate, emphasizing the need for further investigation in this area. Five AMT1 genes (PgAMT1-1 to PgAMT1-5) were identified, all of which contain the PF00909 domain, a feature of ammonium transporters. Various characteristics of these genes, including gene length, coding sequence length, and chromosomal locations, were examined. This study evaluated the isoelectric point, hydropathicity, conserved domains, motifs, and synteny of the PgAMT1 proteins. Phylogenetic analysis confirmed the homology of PgAMT1 genes with previously reported AMT in Arabidopsis and tomato. The tissue-specific expression analysis of PgAMT1 genes revealed distinct patterns: PgAMT1-1 and PgAMT1-2 were predominantly expressed in flowers, PgAMT1-3 exhibited notable expression in roots, leaves, and flowers, PgAMT1-4 was primarily expressed in leaf tissue, while the expression of PgAMT1-5 was detected in both leaves and roots. The impact of salt-induced stress on AMT1 gene expression was also examined, revealing that PgAMT1-1, PgAMT1-2, and PgAMT1-4 expression is reduced under increased salt stress. These expression modifications can help regulate NH4+ assimilation in conditions of elevated salinity, maintaining cellular homeostasis and ion balance. This study contributes to the comprehensive identification of the AMT1s gene family in pomegranate; however, further research on the functional characterization of the identified PgAMT1s is needed.

Integrated transcriptomics and metabolomics analyses provide new insights into cassava in response to nitrogen deficiency

Nitrogen deficiency is a key constraint on crop yield. Cassava, the world’s sixth-largest food crop and a crucial source of feed and industrial materials, can thrive in marginal soils, yet its yield is still significantly affected by limited nitrogen availability. Investigating cassava’s response mechanisms to nitrogen scarcity is therefore essential for advancing molecular breeding and identifying nitrogen-efficient varieties. This research undertook a comprehensive analysis of cassava seedlings’ physiological, gene expression, and metabolite responses under low nitrogen stress. Findings revealed that nitrogen deficiency drastically suppressed seedling growth, significantly reduced nitrate and ammonium transport to aerial parts, and led to a marked increase in carbohydrate, reactive oxygen species, and ammonium ion levels in the leaves. Transcriptomic and metabolomic analyses further demonstrated notable alterations in genes and metabolites linked to carbon and nitrogen metabolism, flavonoid biosynthesis, and the purine metabolic pathway. Additionally, several transcription factors associated with cassava flavonoid biosynthesis under nitrogen-deficient conditions were identified. Overall, this study offers fresh insights and valuable genetic resources for unraveling cassava’s adaptive mechanisms to nitrogen deprivation.

Functional and Molecular Characterization of Plant Nitrate Transporters Belonging to NPF (NRT1/PTR) 6 Subfamily.

Plant nitrate transporters in the NPF (NRT1) family are characterized by multifunctionality and their involvement in a number of physiological processes. The proteins in this family have been identified in many monocotyledonous and dicotyledonous species: a bioinformatic analysis predicts from 20 to 139 members in the plant genomes sequenced so far, including mosses. Plant NPFs are phylogenetically related to proton-coupled oligopeptide transporters, which are evolutionally conserved in all kingdoms of life apart from Archaea. The phylogenetic analysis of the plant NPF family is based on the amino acid sequences present in databases; an analysis identified a separate NPF6 clade (subfamily) with the first plant nitrate transporters studied at the molecular level. The available information proves that proteins of the NPF6 clade play key roles not only in the supply of nitrate and its allocation within different parts of plants but also in the transport of chloride, amino acids, ammonium, and plant hormones such as auxins and ABA. Moreover, members of the NPF6 family participate in the perception of nitrate and ammonium, signaling, plant responses to different abiotic stresses, and the development of tolerance to these stresses and contribute to the structure of the root-soil microbiome composition. The available information allows us to conclude that NPF6 genes are among the promising targets for engineering/editing to increase the productivity of crops and their tolerance to stresses. The present review summarizes the available published data and our own results on members of the NPF6 clade of nitrate transporters, especially under salinity; we outline their molecular, structural, and functional characteristics and suggest potential lines for future research.

Alleviation of drought stress in tomato by foliar application of seafood waste extract

To manage the adverse effects of garbage pollution and avoid using chemicals, a natural extract of seafood shells was obtained and explored for its beneficial role. Physical characterization highlighted that its active compounds correspond to chitin and its derivative, chitosan. The ability of the extracted biostimulant to foster tomato tolerance was tested on drought-stressed plants. Along with changes in morphological parameters, the accumulation of chlorophyll and carotenoids was improved. The biostimulant also mediates the accumulation of osmoprotectants and an increased leaf water content. Furthermore, the biostimulant effectively promotes tolerance by increasing drought-stress SIERF84 Transcription factor and decreasing both SIARF4 and SlWRKY81 transcript levels, which in turn, mediates stomatal closure. In addition, the up-regulation of key genes related to NO3− uptake (NTR1.1/2) and assimilation (NR) coupled with the downregulation of ammonium transporters’ genes (AMT1.1/2), allowed the uptake of NO3− over NH4+ in the tolerant genotype which is likely to be associated with drought tolerance. Overall, the biostimulant was effective in alleviating water stress and showed similar effects to commercial chitosan. Besides the benefits of a circular economy framework, this biostimulant-based approach is innovative to promote a sustainable eco-agriculture, in the face of persistent water scarcity.

The Identification of AMT Family Genes and Their Expression, Function, and Regulation in Chenopodium quinoa.

Quinoa (Chenopodium quinoa) is an Andean allotetraploid pseudocereal crop with higher protein content and balanced amino acid composition in the seeds. Ammonium (NH4+), a direct source of organic nitrogen assimilation, mainly transported by specific transmembrane ammonium transporters (AMTs), plays important roles in the development, yield, and quality of crops. Many AMTs and their functions have been identified in major crops; however, no systematic analyses of AMTs and their regulatory networks, which is important to increase the yield and protein accumulation in the seeds of quinoa, have been performed to date. In this study, the CqAMTs were identified, followed by the quantification of the gene expression, while the regulatory networks were predicted based on weighted gene co-expression network analysis (WGCNA), with the putative transcriptional factors (TFs) having binding sites on the promoters of CqAMTs, nitrate transporters (CqNRTs), and glutamine-synthases (CqGSs), as well as the putative TF expression being correlated with the phenotypes and activities of GSs, glutamate synthase (GOGAT), nitrite reductase (NiR), and nitrate reductase (NR) of quinoa roots. The results showed a total of 12 members of the CqAMT family with varying expressions in different organs and in the same organs at different developmental stages. Complementation expression analyses in the triple mep1/2/3 mutant of yeast showed that except for CqAMT2.2b, 11/12 CqAMTs restored the uptake of NH4+ in the host yeast. CqAMT1.2a was found to mainly locate on the cell membrane, while TFs (e.g., CqNLPs, CqG2Ls, B3 TFs, CqbHLHs, CqZFs, CqMYBs, CqNF-YA/YB/YC, CqNACs, and CqWRKY) were predicted to be predominantly involved in the regulation, transportation, and assimilation of nitrogen. These results provide the functions of CqAMTs and their possible regulatory networks, which will lead to improved nitrogen use efficiency (NUE) in quinoa as well as other major crops.

Molecular characterization and insilico expression analysis of ammonium transporter genes family in Sorghum bicolor

Molecular characterization and insilico expression analysis of ammonium transporter genes family in Sorghum bicolor

Transcriptional and molecular simulation analysis of the response mechanism of anammox bacteria to 3,4-dimethylpyrazole phosphate stress.

Anaerobic ammonium oxidation (anammox) and nitrification are two vital biological pathways for ammonium oxidation, pivotal in microbial nitrogen cycling. 3,4-Dimethylpyrazole phosphate (DMPP) is commonly used as inhibitors in agricultural soils to reduce nitrogen losses from farmland, while whether it affect anammox is an open question. Acute inhibition tests revealed that 53.5 mg·L-1 DMPP caused 50 % reduction in anammox bacteria. After 36 days of prolonged exposure to 5 mg·L-1 DMPP, the ammonium(nitrite) removal rate of endnote decreased from 78.39(94.78) to 13.57(15.28) mgN·gVSS-1·d-1. Additionally, the abundance of Ca. Kuenenia decreased from 36.5 % to 6.06 %. Transcriptomic analysis revealed that the mRNA levels of ammonium transport genes (amt_1 and amt_4), and hydrazine synthase (hzs) were significantly downregulated. Molecular docking simulations indicated that DMPP bound with ammonium transport and hydrazine synthesis. This interaction hindered the transcriptional levels of genes encoding ammonium transporters and hzs. The study has guiding value to reduce the nitrogen loss involved in anammox bacteria in agricultural soils under the application of DMPP.

Cold stress impairs nitrogen uptake and enhances translocation through AMT1 and NRT2 gene regulation in tomato

Nitrogen is a vital nutrient for plant growth, playing a crucial role in various physiological processes. Cold stress significantly impacts plant physiology, including nitrogen uptake and translocation. This study investigates the effects of cold stress on nitrogen dynamics in tomato plants by examining the expression of ammonium (AMT1) and nitrate (NRT2) transporter genes. Under normal conditions, AMT1 and NRT2 genes are predominantly expressed in the roots, with varying levels of expression in other tissues. However, following exposure to cold stress, a significant downregulation of most AMT1 and NRT2 genes in the roots was observed, indicating a reduced capacity for nitrogen uptake and assimilation. Conversely, there was a notable upregulation of these genes in the leaves, suggesting an enhanced capacity for nitrogen translocation and metabolism under cold conditions. This differential expression between roots and leaves highlights the plant's adaptive mechanisms to cope with environmental stress. It indicates a strategy to conserve energy in the roots while increasing nutrient transport in the leaves to support metabolic adjustments. These insights into the molecular basis of nitrogen management under cold stress can inform strategies to enhance crop resilience and productivity.

3D variability analysis reveals a hidden conformational change controlling ammonia transport in human asparagine synthetase

Advances in X-ray crystallography and cryogenic electron microscopy (cryo-EM) offer the promise of elucidating functionally relevant conformational changes that are not easily studied by other biophysical methods. Here we show that 3D variability analysis (3DVA) of the cryo-EM map for wild-type (WT) human asparagine synthetase (ASNS) identifies a functional role for the Arg-142 side chain and test this hypothesis experimentally by characterizing the R142I variant in which Arg-142 is replaced by isoleucine. Support for Arg-142 playing a role in the intramolecular translocation of ammonia between the active site of the enzyme is provided by the glutamine-dependent synthetase activity of the R142 variant relative to WT ASNS, and MD simulations provide a possible molecular mechanism for these findings. Combining 3DVA with MD simulations is a generally applicable approach to generate testable hypotheses of how conformational changes in buried side chains might regulate function in enzymes.

Monitoring ammonia and water transport through anion exchange membranes in direct ammonia fuel cells

Anion exchange membrane direct ammonia fuel cells (AEM-DAFCs) can generate clean electricity with zero carbon emissions. However, their performance are hindered by severe ammonia crossover, which causes a mixed potential at the cathode, thereby reducing the cell voltage. Meanwhile, the produced intermediates, such as Nads species, adsorb onto catalyst, causing poisoning and thus impeding the oxygen reduction reaction (ORR). Another key issue for AEM-DAFCs is to maintain an appropriate water content at the cathode, which is crucial for promoting ORR and avoiding water flooding. In this work, an in-situ observation method is developed to quantitatively characterize the rates of ammonia and water crossover. Besides, a half-cell configuration is designed and developed to evaluate the cathode potential drop caused by ammonia crossover. The effects of operating conditions (such as ammonia concentration and cell temperature) and membrane electrode assembly (MEA) design parameters (such as membrane thickness and cathode wettability) on behaviors of ammonia and water crossover are thoroughly examined. The findings show that ammonia crossover can be substantially reduced by using a low-concentration ammonia solution or a thicker membrane. Moreover, this study offers guidance on developing fuel and water management strategies to enhance the performance of AEM-DAFCs.

OsAMT1.1 knockout-induced decrease in cadmium absorption and accumulation by rice related to cadmium absorption-related gene downregulation

Cadmium (Cd) is a hazardous heavy metal that poses a serious risk to human health through the food chain, with rice being a significant vector because of its tendency to accumulate Cd. Nitrogen (N), an essential element for plant growth, also affects the Cd absorption and accumulation in crops. This study investigated the effects of N application on Cd absorption and accumulation in Cd-contaminated soils. Potting experiment showed that increasing N concentrations significantly increased the plant biomass and Cd contents in rice tissues. Ammonium (NH4+) transporter gene OsAMT1.1 knockout led to a substantial reduction in Cd absorption and accumulation in all rice tissues compared to that in the wild-type plants. Specifically, osamt1.1 mutants increased the Cd content in culm tissues, whereas it was reduced in brown rice. In addition, OsAMT1.1 knockout reduced Cd2+ influx in roots under NH4+-N addition, although OsAMT1.1 lacked Cd transport ability when expressed in yeast. Gene expression analysis revealed that OsAMT1.1 knockout reduced Cd absorption-related genes (OsIRT1, OsNRAMP1, and OsNRAMP5) expression levels. These finding highlight the critical role of N supply and OsAMT1.1 in regulating the Cd content in rice, offering insights into the molecular mechanisms of Cd transportation in plants.

Amine ester improves rice growth and resistance by promoting ammonium and potassium uptake

Fertilizers play a crucial role in improving crop yields; however, excessive fertilizer application leads to environmental pollution, increases greenhouse gas emissions, and contributes to global warming. Therefore, improving fertilizer efficiency is of great significance for crop production and ecological security. Octanoic acid (OA), a type of straight-chain saturated fatty acid commonly found in plants and animals, is known to promote plant growth. In this study, we synthesized amine esters (AE) using OA as the precursor. AE demonstrated a greater growth-promoting effect than OA. To further explore the mechanism underlying AE-induced growth promotion, the responses of macro-element transporter mutants to AE were analyzed. Genetic and physiological studies indicated that mutants of potassium channel AKT1 and ammonium transporter 1 (AMT1) inhibited AE-induced growth promotion in rice seedlings, whereas mutants of nitrate transporter 1.1B (NRT1.1B) and phosphate transporter 8 (PT8) did not significantly inhibit AE-induced growth. Additionally, yeast rescue assays revealed that AE significantly enhanced the absorption of ammonium and potassium ions. Glutamine synthetase 1 (gs1;1) mutants exhibited a response similar to that of AMT1 RNAi, which inhibited AE-induced growth promotion. Furthermore, the administration of AE led to increased chlorophyll accumulation and enhanced resistance to rice blast and sheath blight (ShB) via the potassium and ammonium pathways, respectively. AE also improved tolerance to saline and saline-alkaline stresses through these pathways. In conclusion, AE represents a novel fertilizer additive that promotes rice growth and enhances tolerance to biotic and abiotic stresses by activating ammonium and potassium uptake.

Hydrogen peroxide diffusion across the red blood cell membrane occurs mainly by simple diffusion through the lipid fraction

Hydrogen peroxide (H2O2) is an oxidant produced endogenously by several enzymatic pathways. While it can cause molecular damage, H2O2 also plays a role in regulating cell proliferation and survival through redox signaling pathways. In the vascular system, red blood cells (RBCs) are notably efficient at metabolizing H2O2. In addition to a robust antioxidant defense, we recently determined that human RBCs also have a high membrane permeability to H2O2 that is independent of aquaporin 1 or aquaporin 3. In this work, we sought to further investigate the permeation mechanism of H2O2 through the membrane of human RBCs. First, we explored the role of other erythrocytic membrane proteins in H2O2 transport, including urea transporter B and ammonia transporter Rh proteins. However, no differences were found in H2O2 permeability in RBCs lacking these proteins compared to control RBCs. We then focused on the hypothesis that H2O2 diffuses through the lipid bilayer. To test this, we studied H2O2 permeability in RBCs from patients with Gaucher disease (GD), which accumulate sphingolipids in the membrane, affecting RBC morphology and deformability. We found that RBCs from GD patients exhibited lower H₂O₂ membrane permeability. In another approach, we treated normal RBCs with hexanol, which fluidizes the lipid fraction of the RBC membrane, and observed an increase in the permeability to H2O2. In contrast, hexanol had no effect on the rate of water efflux by aquaporin 1. Together, these results support the hypothesis that H2O2 diffusion through the RBC membrane occurs primarily through the lipid fraction.

Modeling nitrogen recovery and water transport in gas-permeable membranes

This study presents a new modeling approach for nitrogen recovery in gas-permeable membrane (GPM) contactors, including both ammonia and water transport dynamics. A distinct feature of the model is its capacity to model water transport across the membrane, which has been overlooked in most publications. Osmotic pressure differences are used to predict the behavior of ammonia and water transport in the GPM contactor. Experiments carried out to develop, test and calibrate the model examined the dynamics of ammonia and water transport through the GPM contactor at various nitrogen concentrations. Specifically, the GPM contactor was tested for nitrogen recovery from high-strength synthetic wastewaters (2.4–10.6 g N/L) at 35 °C and at pH 9. The initial volume of the trapping solution (diluted H2SO4) was 10 times lower than that of the synthetic wastewater, aiming to concentrate the recovered nitrogen. The estimated ammonia transport constant (Km) ranged between (1.2 - 2.1)·10–6 m/s and water transport constant Kw between (2.8 - 8.2)·10–10 m/(s bar). Numerical determination of the model parameters revealed high R² values, demonstrating strong agreement with experimental data.

Study on the physiological responses and tolerance mechanisms to subchronic carbonate alkalinity exposure in the gills of Paramisgurnus dabryanus

Given the reduction of freshwater resources, saline-alkaline aquaculture has emerged as an effective approach to expand the fishery's accessible space. High carbonate alkalinity (CA) is a major stressor for aquatic organisms in saline-alkaline environments. Paramisgurnus dabryanus is a potential species for culture in saline-alkaline water, making it an ideal model for investigating the physiological responses and tolerance mechanisms to CA exposure in freshwater fishes. In the current study, P. dabryanus were exposed to 15 and 30 mmol/L NaHCO3, combining blood biochemical, gill histological, transcriptomic, and metabolomic methods for conjoint analysis of response mechanisms. After 28-d exposure, the gill ventilation frequency of P. dabryanus decreased significantly, gill lamellae twisted and atrophied, and gill filament epithelial cells proliferated, potentially limiting gas exchange, whereas the accessory air-breathing frequency increased significantly, possibly for greater oxygen uptake. Serum osmolality and blood pH remained relatively steady, while serum ammonia levels rose significantly. A total of 3718 differentially expressed genes (DEGs) and 205 differential metabolites (DMs) were identified between the control group and 30 mmol/L NaHCO3 group, involved in ion transport (Na+/K+-ATPase, V-type ATPase, carbonic anhydrase, and ABC transporters), ammonia transport (Rh glycoproteins and Aquaporins), amino acid metabolism, carbohydrate metabolism, and fatty acid metabolism. Furthermore, DEGs were significantly associated with cell-cell/ extracellular matrix interaction and protein synthesis. An integrated multi-omics analysis revealed the activation of carbon metabolism and TCA cycle. These results indicate that in response to CA exposure, P. dabryanus may facilitate carrier-mediated ion and ammonia transport to maintain the internal osmotic equilibrium and lessen the deleterious effects of blocked ammonia excretion. Meanwhile, amino acid metabolism and protein synthesis are disturbed, P. dabryanus can modulate carbohydrate catabolism to maintain energy homeostasis. The above findings provide novel insights into saline-alkaline adaptation in freshwater fishes, paving the way for future research and development of saline-alkaline-tolerant Cobitidae strains.

Energetic and exergetic analysis of an AE-T100 micro gas turbine system fuelled by partially cracked ammonia

Abstract In recent years, ammonia has gained attention as a fuel for power generation purposes, especially due to its chemical characteristics which ensure reaching the target of zero-carbon emissions. Ammonia is also a powerful hydrogen carrier, with a solid and consolidated production method based on the Haber-Bosh process. By exploiting the ease of transport of ammonia that is liquid at moderately low pressures and temperatures, it can be converted to hydrogen and nitrogen through a cracking reaction directly at the power production plant site. Moreover, to reduce greenhouse gas emissions (GHG) in the production energy sector, small-scale power generation systems are today gaining more attention. In this context, the use of partially cracked ammonia mixtures in micro gas turbine fields represents an interesting solution in the distributed generation sector. In this work, an energetic and exergetic analysis of an AE-T100 mGT fuelled by partially cracked ammonia mixtures is carried out. The analysis has been performed by varying the ammonia cracking percentage according to the maximum achievable conversion by exploiting the thermal energy contained in mGT exhaust gases. The whole system has been modeled in MATLAB/Simulink environment and it includes not only the turbine but also the ammonia conditioning system required to bring the ammonia at the condition imposed by the mGT. Moreover, a comparison with the mGT fuelled by conventional fuel is carried out both from a thermodynamic and exergetic point of view.

Acute ammonia stress affects the immune response, oxidative stress, ammonia transport and detoxication in the hepatopancreas of freshwater mollusk Solenaia oleivora

Ammonia is a common and major pollutant in aquatic systems. Excessive ammonia has toxic effects on hepatopancreas in aquatic animals. In this study, we investigated the toxic effects of acute ammonia (concentration: 10 mg/L; test duration: 48 h) stress on the hepatopancreas of a freshwater mollusk, Solenaia oleivora. Transcriptome analysis identified 3355 differentially expressed genes (DEGs), including 1432 up-regulated and 1923 down-regulated genes. Many DEGs were associated with immune and stress responses, including heat shock proteins, pattern recognition receptors, and lysozyme. In addition, some DEGs were related to ammonia transport and detoxification, such as aquaporins, K+channel, V-ATPase, cytochrome p450, glutathione transferase, and glutamine synthetase. Physiological analysis showed that ammonia stress increased the activities of antioxidant enzymes (superoxide dismutase and catalase) and non-specific immune enzymes (acid phosphatase) and the levels of liver injury markers (malonaldehyde, aspartate aminotransferase, and alanine transaminase). TdT-mediated dUTP nick-end labeling assay revealed that ammonia stress induced apoptosis in the hepatopancreas. These results indicated the toxic effects of ammonia on hepatopancreas on the immune response, oxidative stress, ammonia transport and detoxification of S. oleivora. Our findings will accumulate data on the toxic effects of ammonia on the hepatopancreas of aquatic animals.

Experimental and simulation study on ammonium transport and turbidity changes in a strong leakage zone during surface-water recharge in the Yufu River Basin, China

Experimental and simulation study on ammonium transport and turbidity changes in a strong leakage zone during surface-water recharge in the Yufu River Basin, China

Down-regulation of the rice HRS1 HOMOLOG3 transcriptional repressor gene due to N deficiency directly co-activates ammonium and phosphate transporter genes.

Rice HRS1 HOMOLOG3 (OsHHO3) acts as a transcriptional repressor of AMMONIUM TRANSPORTER1 (OsAMT1) genes in rice; thus, reduced OsHHO3 expression in nitrogen (N)-deficient environments promotes ammonium uptake. In this study, we show that OsHHO3 also functions as a repressor of a specific subset of phosphate (Pi) transporter (PT) genes involved in the uptake and root-to-shoot translocation of Pi, including OsPT2, OsPT4, and OsPHO1;1. Disruption of OsHHO3 increased Pi uptake and Pi contents in shoots and roots, while overexpression of OsHHO3 caused the opposite effects. Furthermore, phosphorus (P) deficiency slightly decreased OsHHO3 expression, up-regulating a specific subset of PT genes. However, N deficiency was more effective than P deficiency in suppressing OsHHO3 expression in roots, and unlike N deficiency-dependent activation of PT genes under the control of OsHHO3, the P deficiency-dependent activation of OsAMT1 genes was minimal. Interestingly, the simultaneous deficiency of both N and P promoted the OsHHO3-regulated expression of PT genes more significantly than the deficiency of either N or P, but diminished the expression of genes regulated by OsPHR2, a master regulator of Pi starvation-responsive transcriptional activation. Phenotypic analysis revealed that the inactivation and overexpression of OsHHO3 improved and reduced plant growth, respectively, under N-deficient and P-deficient conditions. These results indicate that OsHHO3 regulates a specific subset of PT genes independently of OsPHR2-mediated regulation and plays a critical role in the adaptation to diverse N and P environments.

Prokaryotic expression and purification of inorganic nitrogen transporters in wheat

The nitrate transporter (NRT) and ammonium transporter (AMT) are crucial transmembrane proteins involved in the absorption, transport, and distribution of inorganic nitrogen in wheat. Obtaining NRT/AMT and preparing corresponding antibodies are conducive to probing into their tissue localization and comprehending the nitrogen utilization process in wheat. In this study, four genes (TaNPF4.5, TaNPF8.3, TaNRT3.1, TaAMT1.2) with high expression levels were chosen and cloned from 405 genes of the TaNRT/TaNPF family and 23 genes of the TaAMT family identified previously. The transmembrane domains of the four transporters were predicted by HMMER to determine the putative expression segments, followed by prokaryotic expression and purification. Under the induction with 1 mmol/L IPTG at 37 ℃, the non-transmembrane segments of TaNPF4.5, TaNPF8.3, and TaNRT3.1 reached the highest expression levels (as inclusion bodies) after 4 h, while TaAMT1.2 was expressed at the highest level (as a soluble protein) after 3 h. TaNPF4.5, TaNPF8.3, and TaNRT3.1 were purified by a pH gradient. The purity of TaNPF4.5 and TaNPF8.3 reached about 87% and 85% at pH 2.0 and pH 3.0, respectively, both of which were suitable for antibody preparation. However, the purity of TaNRT3.1 did not reach 85%. TaAMT1.2 was purified by an imidazole gradient, reaching the purity of about 95% at 20 mmol/L imidazole, and the antibody was prepared successfully. The expression, purification, and antibody preparation of TaAMT1.2 not only provides insights into the expression, purification, and antibody preparation of membrane proteins including TaNPF4.5 and TaNPF8.3 but also lays a foundation for studying the expression and localization of membrane proteins in wheat.