{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,24]],"date-time":"2026-02-24T20:04:04Z","timestamp":1771963444935,"version":"3.50.1"},"reference-count":59,"publisher":"MDPI AG","issue":"1","license":[{"start":{"date-parts":[[2022,12,20]],"date-time":"2022-12-20T00:00:00Z","timestamp":1671494400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["DL57\/2016\/CP [12345\/2018]\/CT [2475]"],"award-info":[{"award-number":["DL57\/2016\/CP [12345\/2018]\/CT [2475]"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["DL57\/2016\/CP [12345\/2018]\/CT [2477]"],"award-info":[{"award-number":["DL57\/2016\/CP [12345\/2018]\/CT [2477]"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["UIDB\/04046\/2020"],"award-info":[{"award-number":["UIDB\/04046\/2020"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["UIDP\/04046\/2020"],"award-info":[{"award-number":["UIDP\/04046\/2020"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["UIDB\/04129\/2020"],"award-info":[{"award-number":["UIDB\/04129\/2020"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["UIDB\/00329\/2020"],"award-info":[{"award-number":["UIDB\/00329\/2020"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["PTDC\/ASP-AGR\/0760\/2020"],"award-info":[{"award-number":["PTDC\/ASP-AGR\/0760\/2020"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001871","name":"Funda\u00e7\u00e3o para a Ci\u00eancia e Tecnologia","doi-asserted-by":"publisher","award":["PPBI-POCI-01-0145-FEDER-022122"],"award-info":[{"award-number":["PPBI-POCI-01-0145-FEDER-022122"]}],"id":[{"id":"10.13039\/501100001871","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Plants"],"abstract":"<jats:p>In temperate forests, the roots of various tree species are colonized by ectomycorrhizal fungi, which have a key role in the nitrogen nutrition of their hosts. However, not much is known about the molecular mechanisms related to nitrogen metabolism in ectomycorrhizal plants. This study aimed to evaluate the nitrogen metabolic response of oak plants when inoculated with the ectomycorrhizal fungus Pisolithus\u00a0tinctorius. The expression of candidate genes encoding proteins involved in nitrogen uptake and assimilation was investigated in ectomycorrhizal roots. We found that three oak ammonium transporters were over-expressed in root tissues after inoculation, while the expression of amino acid transporters was not modified, suggesting that inorganic nitrogen is the main form of nitrogen transferred by the symbiotic fungus into the roots of the host plant. Analysis by heterologous complementation of a yeast mutant defective in ammonium uptake and GFP subcellular protein localization clearly confirmed that two of these genes encode functional ammonium transporters. Structural similarities between the proteins encoded by these ectomycorrhizal upregulated ammonium transporters, and a well-characterized ammonium transporter from E. coli, suggest a similar transport mechanism, involving deprotonation of NH4+, followed by diffusion of uncharged NH3 into the cytosol. This view is supported by the lack of induction of NH4+ detoxifying mechanisms, such as the GS\/GOGAT pathway, in the oak mycorrhizal roots.<\/jats:p>","DOI":"10.3390\/plants12010010","type":"journal-article","created":{"date-parts":[[2022,12,20]],"date-time":"2022-12-20T04:21:56Z","timestamp":1671510116000},"page":"10","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Nitrogen Acquisition and Transport in the Ectomycorrhizal Symbiosis\u2014Insights from the Interaction between an Oak Tree and Pisolithus tinctorius"],"prefix":"10.3390","volume":"12","author":[{"given":"M\u00f3nica","family":"Sebastiana","sequence":"first","affiliation":[{"name":"BioISI\u2014Instituto de Biosistemas e Ci\u00eancias Integrativas, Faculdade de Ci\u00eancias, Universidade de Lisboa, 1749-016 Lisboa, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8864-0277","authenticated-orcid":false,"given":"Susana","family":"Serrazina","sequence":"additional","affiliation":[{"name":"BioISI\u2014Instituto de Biosistemas e Ci\u00eancias Integrativas, Faculdade de Ci\u00eancias, Universidade de Lisboa, 1749-016 Lisboa, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1505-2140","authenticated-orcid":false,"given":"Filipa","family":"Monteiro","sequence":"additional","affiliation":[{"name":"Linking Landscape, Environment, Agriculture and Food (LEAF), Associated Laboratory TERRA, Instituto Superior de Agronomia (ISA), Universidade de Lisboa, 1349-017 Lisbon, Portugal"},{"name":"Centre for Ecology, Evolution and Environmental Changes (cE3c) & CHANGE\u2014Global Change and Sustainability Institute, Faculdade de Ci\u00eancias da Universidade de Lisboa, Campo Grande, 1749-016 Lisboa, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7197-5612","authenticated-orcid":false,"given":"Daniel","family":"Wipf","sequence":"additional","affiliation":[{"name":"Agro\u00e9cologie, INRAE, Institut Agro, University Bourgogne Franche-Comt\u00e9, F-21000 Dijon, France"}]},{"given":"J\u00e9rome","family":"Fromentin","sequence":"additional","affiliation":[{"name":"Agro\u00e9cologie, INRAE, Institut Agro, University Bourgogne Franche-Comt\u00e9, F-21000 Dijon, France"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9373-0760","authenticated-orcid":false,"given":"Rita","family":"Teixeira","sequence":"additional","affiliation":[{"name":"BioISI\u2014Instituto de Biosistemas e Ci\u00eancias Integrativas, Faculdade de Ci\u00eancias, Universidade de Lisboa, 1749-016 Lisboa, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5287-869X","authenticated-orcid":false,"given":"Rui","family":"Malh\u00f3","sequence":"additional","affiliation":[{"name":"BioISI\u2014Instituto de Biosistemas e Ci\u00eancias Integrativas, Faculdade de Ci\u00eancias, Universidade de Lisboa, 1749-016 Lisboa, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2789-7818","authenticated-orcid":false,"given":"Pierre-Emmanuel","family":"Courty","sequence":"additional","affiliation":[{"name":"Agro\u00e9cologie, INRAE, Institut Agro, University Bourgogne Franche-Comt\u00e9, F-21000 Dijon, France"}]}],"member":"1968","published-online":{"date-parts":[[2022,12,20]]},"reference":[{"key":"ref_1","unstructured":"Smith, S.E., and Read, D. (2008). Mycorrhizal Symbiosis, Elsevier. [3rd ed.]."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1023\/A:1020221528515","article-title":"Contribution of Amino Compounds to Dissolved Organic Nitrogen in Forest Soils","volume":"61","author":"Yu","year":"2002","journal-title":"Biogeochemistry"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"473","DOI":"10.1017\/S1464793103006316","article-title":"Diversity and Classification of Mycorrhizal Associations","volume":"79","author":"Brundrett","year":"2004","journal-title":"Biol. Rev. Camb. Philos. Soc."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1658","DOI":"10.3389\/fpls.2019.01658","article-title":"Digging Deeper: In Search of the Mechanisms of Carbon and Nitrogen Exchange in Ectomycorrhizal Symbioses","volume":"10","author":"Stuart","year":"2020","journal-title":"Front. Plant Sci."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"4","DOI":"10.1080\/07352689.2014.897897","article-title":"Inorganic Nitrogen Uptake and Transport in Beneficial Plant Root-Microbe Interactions","volume":"34","author":"Courty","year":"2015","journal-title":"CRC. Crit. Rev. Plant Sci."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"105","DOI":"10.1111\/j.1469-8137.1992.tb01063.x","article-title":"Utilization of Organic and Inorganic Nitrogen Sources by Ectomycorrhizal Fungi in Pure Culture and in Symbiosis with Pinus Contorta Dougl. Ex Loud","volume":"120","author":"Finlay","year":"1992","journal-title":"New Phytol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"819","DOI":"10.1038\/nature03610","article-title":"Nitrogen Transfer in the Arbuscular Mycorrhizal Symbiosis","volume":"435","author":"Govindarajulu","year":"2005","journal-title":"Nature"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"199","DOI":"10.1111\/j.1469-8137.2008.02630.x","article-title":"Arbuscular Mycorrhizal Fungi Can Transfer Substantial Amounts of Nitrogen to Their Host Plant from Organic Material","volume":"181","author":"Leigh","year":"2009","journal-title":"New Phytol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"263","DOI":"10.1016\/j.tplants.2006.04.005","article-title":"Ammonia: A Candidate for Nitrogen Transfer at the Mycorrhizal Interface","volume":"11","author":"Chalot","year":"2006","journal-title":"Trends Plant Sci."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"137","DOI":"10.1111\/j.1469-8137.2007.01992.x","article-title":"The Expanded Family of Ammonium Transporters in the Perennial Poplar Plant","volume":"174","author":"Couturier","year":"2007","journal-title":"New Phytol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1016\/j.phytochem.2006.09.021","article-title":"Nitrogen Transport in the Ectomycorrhiza Association: The Hebeloma Cylindrosporum-Pinus Pinaster Model","volume":"68","author":"Avolio","year":"2007","journal-title":"Phytochemistry"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"735","DOI":"10.1007\/s00572-020-00983-7","article-title":"New Insights into HcPTR2A and HcPTR2B, Two High-Affinity Peptide Transporters from the Ectomycorrhizal Model Fungus Hebeloma Cylindrosporum","volume":"30","author":"Ludewig","year":"2020","journal-title":"Mycorrhiza"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Larsen, P.E., Sreedasyam, A., Trivedi, G., Podila, G.K., Cseke, L.J., and Collart, F.R. (2011). Using next Generation Transcriptome Sequencing to Predict an Ectomycorrhizal Metabolome. BMC Syst. Biol., 5.","DOI":"10.1186\/1752-0509-5-70"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"2666","DOI":"10.1038\/ismej.2017.116","article-title":"Role of Plant-Fungal Nutrient Trading and Host Control in Determining the Competitive Success of Ectomycorrhizal Fungi","volume":"11","author":"Hortal","year":"2017","journal-title":"ISME J."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"914","DOI":"10.1038\/31921","article-title":"Boreal Forest Plants Take up Organic Nitrogen","volume":"392","author":"Ekblad","year":"1998","journal-title":"Nature"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1047","DOI":"10.1111\/nph.15257","article-title":"Nitrogen and Phosphate Metabolism in Ectomycorrhizas","volume":"220","author":"Nehls","year":"2018","journal-title":"New Phytol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"697","DOI":"10.1111\/j.1469-8137.2005.01535.x","article-title":"The High-Affinity Poplar Ammonium Importer PttAMT1.2 and Its Role in Ectomycorrhizal Symbiosis","volume":"168","author":"Selle","year":"2005","journal-title":"New Phytol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"853","DOI":"10.1111\/nph.12199","article-title":"The Family of Ammonium Transporters (AMT) in Sorghum bicolor: Two AMT Members Are Induced Locally, but Not Systemically in Roots Colonized by Arbuscular Mycorrhizal Fungi","volume":"198","author":"Koegel","year":"2013","journal-title":"New Phytol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/j.plaphy.2013.11.029","article-title":"Transcriptional Regulation of Host NH4+ Transporters and GS\/GOGAT Pathway in Arbuscular Mycorrhizal Rice Roots","volume":"75","author":"Ferrol","year":"2014","journal-title":"Plant Physiol. Biochem."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1411","DOI":"10.1093\/pcp\/pcq099","article-title":"Localized Expression of Arbuscular Mycorrhiza-Inducible Ammonium Transporters in Soybean","volume":"51","author":"Kobae","year":"2010","journal-title":"Plant Cell Physiol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"73","DOI":"10.1104\/pp.109.136390","article-title":"A Mycorrhizal-Specific Ammonium Transporter from Lotus japonicus Acquires Nitrogen Released by Arbuscular Mycorrhizal Fungi","volume":"150","author":"Guether","year":"2009","journal-title":"Plant Physiol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1352","DOI":"10.1105\/tpc.114.131144","article-title":"Suppression of Arbuscule Degeneration in Medicago truncatula Phosphate Transporter4 Mutants Is Dependent on the Ammonium Transporter 2 Family Protein AMT2;3","volume":"27","author":"Floss","year":"2015","journal-title":"Plant Cell"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1669","DOI":"10.1111\/pce.12692","article-title":"Deciphering the Molecular Basis of Ammonium Uptake and Transport in Maritime Pine","volume":"39","author":"Fan","year":"2016","journal-title":"Plant Cell Environ."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1141","DOI":"10.1093\/aob\/mcq028","article-title":"Nitrogen Uptake, Assimilation and Remobilization in Plants: Challenges for Sustainable and Productive Agriculture","volume":"105","author":"Dechorgnat","year":"2010","journal-title":"Ann. Bot."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"378","DOI":"10.1046\/j.1365-2958.2000.01704.x","article-title":"Cross-Talk between Ammonium Transporters in Yeast and Interference by the Soybean SAT1 Protein","volume":"35","author":"Marini","year":"2000","journal-title":"Mol. Microbiol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"331","DOI":"10.1046\/j.1365-2958.2000.01994.x","article-title":"Membrane Topology of the Mep\/Amt Family of Ammonium Transporters","volume":"37","author":"Thomas","year":"2000","journal-title":"Mol. Microbiol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"695","DOI":"10.1007\/s00572-017-0786-8","article-title":"Phylogenetic, Structural, and Functional Characterization of AMT3;1, an Ammonium Transporter Induced by Mycorrhization among Model Grasses","volume":"27","author":"Koegel","year":"2017","journal-title":"Mycorrhiza"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"177","DOI":"10.1139\/g02-121","article-title":"An Expression CDNA Library for Suppression Cloning in Yeast Mutants, Complementation of a Yeast His4 Mutant, and EST Analysis from the Symbiotic Basidiomycete Hebeloma Cylindrosporum","volume":"46","author":"Wipf","year":"2003","journal-title":"Genome"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"4282","DOI":"10.1128\/MCB.17.8.4282","article-title":"A Family of Ammonium Transporters in Saccharomyces Cerevisiae","volume":"17","author":"Marini","year":"1997","journal-title":"Mol. Cell. Biol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2012","DOI":"10.1371\/journal.pbio.0040351","article-title":"Ammonium Toxicity and Potassium Limitation in Yeast","volume":"4","author":"Hess","year":"2006","journal-title":"PLoS Biol."},{"key":"ref_31","first-page":"1587","article-title":"Mechanism of Ammonia Transport by Amt\/MEP\/Rh: Structure of AmtB at 1.35 \u00c5","volume":"305","author":"Khademi","year":"2004","journal-title":"New Ser."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"von Wittgenstein, N.J., Le, C.H., Hawkins, B.J., and Ehlting, J. (2014). Evolutionary Classification of Ammonium, Nitrate, and Peptide Transporters in Land Plants. BMC Evol. Biol., 14.","DOI":"10.1186\/1471-2148-14-11"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"679","DOI":"10.3389\/fpls.2016.00679","article-title":"GintAMT3-a Low-Affinity Ammonium Transporter of the Arbuscular Mycorrhizal Rhizophagus Irregularis","volume":"7","author":"Calabrese","year":"2016","journal-title":"Front. Plant Sci."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"7082","DOI":"10.1038\/s41598-018-25430-8","article-title":"Phylogenetic Analyses and In-Seedling Expression of Ammonium and Nitrate Transporters in Wheat","volume":"8","author":"Bajgain","year":"2018","journal-title":"Sci. Rep."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"10419","DOI":"10.1021\/ja300129x","article-title":"Ammonium Transporters Achieve Charge Transfer by Fragmenting Their Substrate","volume":"134","author":"Wang","year":"2012","journal-title":"J. Am. Chem. Soc."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"102","DOI":"10.1016\/j.fgb.2005.10.005","article-title":"GintAMT1 Encodes a Functional High-Affinity Ammonium Transporter That Is Expressed in the Extraradical Mycelium of Glomus Intraradices","volume":"43","author":"Valderas","year":"2006","journal-title":"Fungal Genet. Biol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"247","DOI":"10.1007\/s00572-018-0823-2","article-title":"Ectomycorrhizal Inoculation with Pisolithus tinctorius Reduces Stress Induced by Drought in Cork Oak","volume":"28","author":"Sebastiana","year":"2018","journal-title":"Mycorrhiza"},{"key":"ref_38","first-page":"197","article-title":"Comparing Total Nitrogen And Crude Protein Content Of Green Coffee Beans (Coffea Spp.) From Different Geographical Origins","volume":"5","author":"Rodrigues","year":"2010","journal-title":"Coffe Sci."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"180069","DOI":"10.1038\/sdata.2018.69","article-title":"The Draft Genome Sequence of Cork Oak","volume":"5","author":"Ramos","year":"2018","journal-title":"Sci. Data"},{"key":"ref_40","unstructured":"(2021, March 20). The Arabidopsis Information Resource. Available online: https:\/\/www.arabidopsis.org\/."},{"key":"ref_41","unstructured":"(2021, March 20). The Pant Genomics Resource, Available online: https:\/\/phytozome-next.jgi.doe.gov\/."},{"key":"ref_42","unstructured":"(2021, May 15). Pfam. Available online: https:\/\/pfam.xfam.org\/."},{"key":"ref_43","unstructured":"(2021, May 15). SignalP. Available online: https:\/\/services.healthtech.dtu.dk\/service.php?SignalP."},{"key":"ref_44","unstructured":"(2021, May 15). DeepTMHMM. Available online: https:\/\/dtu.biolib.com\/DeepTMHMM\/."},{"key":"ref_45","unstructured":"(2021, May 15). TargetP. Available online: https:\/\/services.healthtech.dtu.dk\/service.php?TargetP-2.0."},{"key":"ref_46","unstructured":"(2021, December 10). Multiple Sequence Alignment by CLUSTALW. Available online: https:\/\/www.genome.jp\/tools-bin\/clustalw."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"1160","DOI":"10.1093\/bib\/bbx108","article-title":"MAFFT Online Service: Multiple Sequence Alignment, Interactive Sequence Choice and Visualization","volume":"20","author":"Katoh","year":"2018","journal-title":"Brief. Bioinform."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"1972","DOI":"10.1093\/bioinformatics\/btp348","article-title":"TrimAl: A Tool for Automated Alignment Trimming in Large-Scale Phylogenetic Analyses","volume":"25","year":"2009","journal-title":"Bioinformatics"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"1596","DOI":"10.1093\/molbev\/msm092","article-title":"MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) Software Version 4.0","volume":"24","author":"Tamura","year":"2007","journal-title":"Mol. Biol. Evol."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"1547","DOI":"10.1093\/molbev\/msy096","article-title":"MEGA X: Molecular Evolutionary Genetics Analysis across Computing Platforms","volume":"35","author":"Kumar","year":"2018","journal-title":"Mol. Biol. Evol."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1307","DOI":"10.1093\/molbev\/msn067","article-title":"An Improved General Amino Acid Replacement Matrix","volume":"25","author":"Le","year":"2008","journal-title":"Mol. Biol. Evol."},{"key":"ref_52","unstructured":"(2022, November 28). FigTree. Available online: http:\/\/tree.bio.ed.ac.uk\/software\/figtree\/."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"7","DOI":"10.1006\/abio.1994.1538","article-title":"A Modified Hot Borate Method Significantly Enhances the Yield of High-Quality RNA from Cotton (Gossypium Hirsutum L.)","volume":"223","author":"Wan","year":"1994","journal-title":"Anal. Biochem."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"157","DOI":"10.1016\/j.plaphy.2019.09.032","article-title":"The Leaf Lipid Composition of Ectomycorrhizal Oak Plants Shows a Drought-Tolerance Signature","volume":"144","author":"Sebastiana","year":"2019","journal-title":"Plant Physiol. Biochem."},{"key":"ref_55","unstructured":"(2021, November 05). Primer-BLAST, Available online: https:\/\/www.ncbi.nlm.nih.gov\/tools\/primer-blast\/."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"1101","DOI":"10.1038\/nprot.2008.73","article-title":"Analyzing Real-Time PCR Data by the Comparative CT Method","volume":"3","author":"Schmittgen","year":"2008","journal-title":"Nat. Protoc."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"691","DOI":"10.1002\/yea.320070704","article-title":"An Efficient Transformation Procedure Enabling Long-Term Storage of Competent Cells of Various Yeast Genera","volume":"7","author":"Dohmen","year":"1991","journal-title":"Yeast"},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"2019","DOI":"10.1038\/nprot.2006.286","article-title":"Rapid, Transient Expression of Fluorescent Fusion Proteins in Tobacco Plants and Generation of Stably Transformed Plants","volume":"1","author":"Sparkes","year":"2006","journal-title":"Nat. Protoc."},{"key":"ref_59","unstructured":"(2022, February 12). SWISS-MODEL. Available online: https:\/\/swissmodel.expasy.org\/."}],"container-title":["Plants"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2223-7747\/12\/1\/10\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T01:44:32Z","timestamp":1760147072000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2223-7747\/12\/1\/10"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,12,20]]},"references-count":59,"journal-issue":{"issue":"1","published-online":{"date-parts":[[2023,1]]}},"alternative-id":["plants12010010"],"URL":"https:\/\/doi.org\/10.3390\/plants12010010","relation":{},"ISSN":["2223-7747"],"issn-type":[{"value":"2223-7747","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,12,20]]}}}