Commonly studied in the context of legume-rhizobia symbiosis, biological nitrogen fixation (BNF) is a key component of the nitrogen cycle in nature. Despite its potential in plant breeding and many years of research, information is still lacking as to the regulation of hundreds of genes connected with plant-bacteria interaction, nodulation, and nitrogen fixation. Here, we compared root nodule transcriptomes of red clover (Trifolium pratense L.) genotypes with contrasting nitrogen fixation efficiency, and we found 491 differentially expressed genes (DEGs) between plants with high and low BNF efficiency. The annotation of genes expressed in nodules revealed more than 800 genes not yet experimentally confirmed. Among genes mediating nodule development, four nod-ule-specific cysteine-rich (NCR) peptides were confirmed in the nodule transcriptome. Gene duplication analyses revealed that genes originating from tandem and dispersed duplication are significantly over-represented among DEGs. Weighted correlation network analysis (WGCNA) organized expression profiles of the transcripts into 16 modules linked to the analyzed traits, such as nitrogen fixation efficiency or sample-specific modules. Overall, the results obtained broaden our knowledge about transcriptomic landscapes of red clover's root nodules and shift the phenotypic description of BNF efficiency on the level of gene expression in situ.

Agricultural Research Ltd Zahradní 1 664 41 Troubsko Czech Republic

Department of Experimental Biology Faculty of Sciences Masaryk University 611 37 Brno Czech Republic

Zobrazit více v PubMed

Lewis G.P., Schrire B.D., Mackinder B.A., Rico L., Clark R. A 2013 Linear Sequence of Legume Genera Set in a Phylogenetic Context—A Tool for Collections Management and Taxon Sampling. South Afr. J. Bot. 2013;89:76–84. doi: 10.1016/j.sajb.2013.06.005. DOI

Azani N., Babineau M., Bailey C.D., Banks H., Barbosa A.R., Pinto R.B., Boatwright J.S., Borges L.M., Brown G.K., Bruneau A., et al. A New Subfamily Classification of the Leguminosae Based on a Taxonomically Comprehensive Phylogeny—The Legume Phylogeny Working Group (LPWG) Taxon. 2017;66:44–77. doi: 10.12705/661.3. DOI

Bell C.J. The Medicago Genome Initiative: A Model Legume Database. Nucleic Acids Res. 2001;29:114–117. doi: 10.1093/nar/29.1.114. PubMed DOI PMC

Udvardi M.K., Tabata S., Parniske M., Stougaard J. Lotus Japonicus: Legume Research in the Fast Lane. Trends Plant Sci. 2005;10:222–228. doi: 10.1016/j.tplants.2005.03.008. PubMed DOI

Rose R.J. Medicago Truncatula as a Model for Understanding Plant Interactions with Other Organisms, Plant Development and Stress Biology: Past, Present and Future. Funct. Plant Biol. 2008;35:253. doi: 10.1071/FP07297. PubMed DOI

Crews T.E. New Perspectives on Nitrogen Cycling in the Temperate and Tropical Americas. Springer; Dordrecht, The Netherlands: 1999. The Presence of Nitrogen Fixing Legumes in Terrestrial Communities: Evolutionary vs Ecological Considerations; pp. 233–246.

Zohary M., Heller D. The Genus Trifolium. Israel Academy of Sciences and Humanities; Jerusalem, Israel: 1984.

Yilmaz A., Yeltekin Y. The Evaluations Of Taxonomic Classifications In The Genus Trifolium L. Based On ITS Sequences. Sak. Univ. J. Sci. 2022;26:545–553. doi: 10.16984/saufenbilder.1074625. DOI

Kintl A., Elbl J., Lošák T., Vaverková M., Nedělník J. Mixed Intercropping of Wheat and White Clover to Enhance the Sustainability of the Conventional Cropping System: Effects on Biomass Production and Leaching of Mineral Nitrogen. Sustainability. 2018;10:3367. doi: 10.3390/su10103367. DOI

Hidalgo L.A., Chedraui P.A., Morocho N., Ross S., San Miguel G. The Effect of Red Clover Isoflavones on Menopausal Symptoms, Lipids and Vaginal Cytology in Menopausal Women: A Randomized, Double-Blind, Placebo-Controlled Study. Gynecol. Endocrinol. 2005;21:257–264. doi: 10.1080/09513590500361192. PubMed DOI

Occhiuto F., Pasquale R.D., Guglielmo G., Palumbo D.R., Zangla G., Samperi S., Renzo A., Circosta C. Effects of Phytoestrogenic Isoflavones from Red Clover (Trifolium pratense L.) on Experimental Osteoporosis. Phytother. Res. 2007;21:130–134. doi: 10.1002/ptr.2037. PubMed DOI

Akbaribazm M., Khazaei F., Naseri L., Pazhouhi M., Zamanian M., Khazaei M. Pharmacological and Therapeutic Properties of the Red Clover (Trifolium pratense L.): An Overview of the New Findings. J. Tradit. Chin. Med. 2021;41:8. PubMed

Trněný O., Vlk D., Macková E., Matoušková M., Řepková J., Nedělník J., Hofbauer J., Vejražka K., Jakešová H., Jansa J., et al. Allelic Variants for Candidate Nitrogen Fixation Genes Revealed by Sequencing in Red Clover (Trifolium pratense L.) Int. J. Mol. Sci. 2019;20:5470. doi: 10.3390/ijms20215470. PubMed DOI PMC

Kouchi H., Imaizumi-Anraku H., Hayashi M., Hakoyama T., Nakagawa T., Umehara Y., Suganuma N., Kawaguchi M. How Many Peas in a Pod? Legume Genes Responsible for Mutualistic Symbioses Underground. Plant Cell Physiol. 2010;51:1381–1397. doi: 10.1093/pcp/pcq107. PubMed DOI PMC

Roux B., Rodde N., Jardinaud M.-F., Timmers T., Sauviac L., Cottret L., Carrère S., Sallet E., Courcelle E., Moreau S., et al. An Integrated Analysis of Plant and Bacterial Gene Expression in Symbiotic Root Nodules Using Laser-Capture Microdissection Coupled to RNA Sequencing. Plant J. 2014;77:817–837. doi: 10.1111/tpj.12442. PubMed DOI

Gage D.J. Infection and Invasion of Roots by Symbiotic, Nitrogen-Fixing Rhizobia during Nodulation of Temperate Legumes. Microbiol. Mol. Biol. Rev. 2004;68:280–300. doi: 10.1128/MMBR.68.2.280-300.2004. PubMed DOI PMC

Masson-Boivin C., Giraud E., Perret X., Batut J. Establishing Nitrogen-Fixing Symbiosis with Legumes: How Many Rhizobium Recipes? Trends Microbiol. 2009;17:458–466. doi: 10.1016/j.tim.2009.07.004. PubMed DOI

Zhang X.-X., Turner S.L., Guo X.-W., Yang H.-J., Debellé F., Yang G.-P., Dénarié J., Young J.P.W., Li F.-D. The Common Nodulation Genes of Astragalus Sinicus Rhizobia Are Conserved despite Chromosomal Diversity. Appl. Environ. Microbiol. 2000;66:2988–2995. doi: 10.1128/AEM.66.7.2988-2995.2000. PubMed DOI PMC

Bek A.S., Sauer J., Thygesen M.B., Duus J.Ø., Petersen B.O., Thirup S., James E., Jensen K.J., Stougaard J., Radutoiu S. Improved Characterization of Nod Factors and Genetically Based Variation in LysM Receptor Domains Identify Amino Acids Expendable for Nod Factor Recognition in Lotus Spp. Mol. Plant. Microbe Interact. 2010;23:58–66. doi: 10.1094/MPMI-23-1-0058. PubMed DOI

Riely B.K., Ané J.-M., Penmetsa R.V., Cook D.R. Genetic and Genomic Analysis in Model Legumes Bring Nod-Factor Signaling to Center Stage. Curr. Opin. Plant Biol. 2004;7:408–413. doi: 10.1016/j.pbi.2004.04.005. PubMed DOI

Kassaw T., Nowak S., Schnabel E., Frugoli J. ROOT DETERMINED NODULATION1 Is Required for M. Truncatula CLE12, But Not CLE13, Peptide Signaling through the SUNN Receptor Kinase. Plant Physiol. 2017;174:2445–2456. doi: 10.1104/pp.17.00278. PubMed DOI PMC

Gleason C., Chaudhuri S., Yang T., Munoz A., Poovaiah B., Oldroyd G.E. Nodulation Independent of Rhizobia Induced by a Calcium-Activated Kinase Lacking Autoinhibition. Nature. 2006;441:1149–1152. doi: 10.1038/nature04812. PubMed DOI

Tirichine L., Sandal N., Madsen L.H., Radutoiu S., Albrektsen A.S., Sato S., Asamizu E., Tabata S., Stougaard J. A Gain-of-Function Mutation in a Cytokinin Receptor Triggers Spontaneous Root Nodule Organogenesis. Science. 2007;315:104–107. doi: 10.1126/science.1132397. PubMed DOI

Schultze M., Kondorosi A. Regulation of Symbiotic Root Nodule Development. Annu. Rev. Genet. 1998;32:33. doi: 10.1146/annurev.genet.32.1.33. PubMed DOI

Downie J.A. Legume Nodulation. Curr. Biol. 2014;24:R184–R190. doi: 10.1016/j.cub.2014.01.028. PubMed DOI

Brill W.J. Biochemical Genetics of Nitrogen Fixation. Microbiol. Rev. 1980;44:449–467. doi: 10.1128/mr.44.3.449-467.1980. PubMed DOI PMC

Ott T., van Dongen J.T., Günther C., Krusell L., Desbrosses G., Vigeolas H., Bock V., Czechowski T., Geigenberger P., Udvardi M.K. Symbiotic Leghemoglobins Are Crucial for Nitrogen Fixation in Legume Root Nodules but Not for General Plant Growth and Development. Curr. Biol. 2005;15:531–535. doi: 10.1016/j.cub.2005.01.042. PubMed DOI

Nap J.-P., Bisseling T. Molecular Biology of Symbiotic Nitrogen Fixation. CRC Press; Boca Raton, FL, USA: 2018. Nodulin Function and Nodulin Gene Regulation in Root Nodule Development; pp. 181–229.

Mergaert P., Uchiumi T., Alunni B., Evanno G., Cheron A., Catrice O., Mausset A.-E., Barloy-Hubler F., Galibert F., Kondorosi A., et al. Eukaryotic Control on Bacterial Cell Cycle and Differentiation in the Rhizobium –Legume Symbiosis. Proc. Natl. Acad. Sci. USA. 2006;103:5230–5235. doi: 10.1073/pnas.0600912103. PubMed DOI PMC

Haag A.F., Arnold M.F.F., Myka K.K., Kerscher B., Dall’Angelo S., Zanda M., Mergaert P., Ferguson G.P. Molecular Insights into Bacteroid Development during Rhizobium– Legume Symbiosis. FEMS Microbiol. Rev. 2013;37:364–383. doi: 10.1111/1574-6976.12003. PubMed DOI

Alunni B., Gourion B. Terminal Bacteroid Differentiation in the Legume−Rhizobium Symbiosis: Nodule-Specific Cysteine-Rich Peptides and Beyond. New Phytol. 2016;211:411–417. doi: 10.1111/nph.14025. PubMed DOI

Montiel J., Downie J.A., Farkas A., Bihari P., Herczeg R., Bálint B., Mergaert P., Kereszt A., Kondorosi É. Morphotype of Bacteroids in Different Legumes Correlates with the Number and Type of Symbiotic NCR Peptides. Proc. Natl. Acad. Sci. USA. 2017;114:5041–5046. doi: 10.1073/pnas.1704217114. PubMed DOI PMC

Zhou P., Silverstein K.A., Gao L., Walton J.D., Nallu S., Guhlin J., Young N.D. Detecting Small Plant Peptides Using SPADA (Small Peptide Alignment Discovery Application) BMC Bioinform. 2013;14:335. doi: 10.1186/1471-2105-14-335. PubMed DOI PMC

Downie J.A., Kondorosi E. Why Should Nodule Cysteine-Rich (NCR) Peptides Be Absent From Nodules of Some Groups of Legumes but Essential for Symbiotic N-Fixation in Others? Front. Agron. 2021;3:654576. doi: 10.3389/fagro.2021.654576. DOI

Roy S., Liu W., Nandety R.S., Crook A., Mysore K.S., Pislariu C.I., Frugoli J., Dickstein R., Udvardi M.K. Celebrating 20 Years of Genetic Discoveries in Legume Nodulation and Symbiotic Nitrogen Fixation. Plant Cell. 2020;32:15–41. doi: 10.1105/tpc.19.00279. PubMed DOI PMC

Sagan M., Morandi D., Tarenghi E., Duc G. Selection of Nodulation and Mycorrhizal Mutants in the Model Plant Medicago Truncatula (Gaertn.) after γ-Ray Mutagenesis. Plant Sci. 1995;111:63–71. doi: 10.1016/0168-9452(95)04229-N. DOI

Penmetsa R.V., Cook D.R. Production and Characterization of Diverse Developmental Mutants of Medicago Truncatula. Plant Physiol. 2000;123:1387–1398. doi: 10.1104/pp.123.4.1387. PubMed DOI PMC

Szczyglowski K., Shaw R.S., Wopereis J., Copeland S., Hamburger D., Kasiborski B., Dazzo F.B., de Bruijn F.J. Nodule Organogenesis and Symbiotic Mutants of the Model Legume Lotus Japonicus. Mol. Plant-Microbe Interact.®. 1998;11:684–697. doi: 10.1094/MPMI.1998.11.7.684. DOI

Bolon Y.-T., Haun W.J., Xu W.W., Grant D., Stacey M.G., Nelson R.T., Gerhardt D.J., Jeddeloh J.A., Stacey G., Muehlbauer G.J., et al. Phenotypic and Genomic Analyses of a Fast Neutron Mutant Population Resource in Soybean. Plant Physiol. 2011;156:240–253. doi: 10.1104/pp.110.170811. PubMed DOI PMC

Thykjaer T., Stiller J., Handberg K., Jones J., Stougaard J. The Maize Transposable Element Ac Is Mobile in the Legume Lotus Japonicus. Plant Mol. Biol. 1995;27:981–993. doi: 10.1007/BF00037025. PubMed DOI

Schauser L., Handberg K., Sandal N., Stiller J., Thykjaer T., Pajuelo E., Nielsen A., Stougaard J. Symbiotic Mutants Deficient in Nodule Establishment Identified after T-DNA Transformation of Lotus Japonicus. Mol. Gen. Genet. MGG. 1998;259:414–423. doi: 10.1007/s004380050831. PubMed DOI

Scholte M., d’Erfurth I., Rippa S., Mondy S., Cosson V., Durand P., Breda C., Trinh H., Rodriguez-Llorente I., Kondorosi E. T-DNA Tagging in the Model Legume Medicago Truncatula Allows Efficient Gene Discovery. Mol. Breed. 2002;10:203–215. doi: 10.1023/A:1020564612093. DOI

Tadege M., Wen J., He J., Tu H., Kwak Y., Eschstruth A., Cayrel A., Endre G., Zhao P.X., Chabaud M., et al. Large-Scale Insertional Mutagenesis Using the Tnt1 Retrotransposon in the Model Legume Medicago truncatula. Plant J. 2008;54:335–347. doi: 10.1111/j.1365-313X.2008.03418.x. PubMed DOI

Pislariu C.I., Murray J.D., Wen J., Cosson V., Muni R.R.D., Wang M., Benedito V.A., Andriankaja A., Cheng X., Jerez I.T., et al. A Medicago truncatula Tobacco Retrotransposon Insertion Mutant Collection with Defects in Nodule Development and Symbiotic Nitrogen Fixation. Plant Physiol. 2012;159:1686–1699. doi: 10.1104/pp.112.197061. PubMed DOI PMC

Fukai E., Stougaard J., Hayashi M. Activation of an Endogenous Retrotransposon Associated with Epigenetic Changes in Lotus Japonicus: A Tool for Functional Genomics in Legumes. Plant Genome. 2013;6:1–11. doi: 10.3835/plantgenome2013.04.0009. DOI

Małolepszy A., Mun T., Sandal N., Gupta V., Dubin M., Urbański D., Shah N., Bachmann A., Fukai E., Hirakawa H., et al. The LORE 1 Insertion Mutant Resource. Plant J. 2016;88:306–317. doi: 10.1111/tpj.13243. PubMed DOI

Cordoba E., Shishkova S., Vance C.P., Hernández G. Antisense Inhibition of NADH Glutamate Synthase Impairs Carbon/Nitrogen Assimilation in Nodules of Alfalfa (Medicago sativa L.): Antisense NADH-GOGAT in Alfalfa Nodules. Plant J. 2003;33:1037–1049. doi: 10.1046/j.1365-313X.2003.01686.x. PubMed DOI

Kumagai H., Kouchi H. Gene Silencing by Expression of Hairpin RNA in Lotus japonicus Roots and Root Nodules. Mol. Plant-Microbe Interactions®. 2003;16:663–668. doi: 10.1094/MPMI.2003.16.8.663. PubMed DOI

Arthikala M.-K., Nanjareddy K., Lara M. RNA Interference. Avid Science; Berlin, Germany: 2017. RNA Interference: A Powerful Functional Analysis Tool for Studying Legume Symbioses; pp. 1–45.

Jacobs T.B., LaFayette P.R., Schmitz R.J., Parrott W.A. Targeted Genome Modifications in Soybean with CRISPR/Cas9. BMC Biotechnol. 2015;15:16. doi: 10.1186/s12896-015-0131-2. PubMed DOI PMC

Wang L., Wang L., Tan Q., Fan Q., Zhu H., Hong Z., Zhang Z., Duanmu D. Efficient Inactivation of Symbiotic Nitrogen Fixation Related Genes in Lotus japonicus Using CRISPR-Cas9. Front. Plant Sci. 2016;7:1333. doi: 10.3389/fpls.2016.01333. PubMed DOI PMC

Curtin S.J., Tiffin P., Guhlin J., Trujillo D.I., Burghardt L.T., Atkins P., Baltes N.J., Denny R., Voytas D.F., Stupar R.M., et al. Validating Genome-Wide Association Candidates Controlling Quantitative Variation in Nodulation. Plant Physiol. 2017;173:921–931. doi: 10.1104/pp.16.01923. PubMed DOI PMC

Badhan S., Ball A.S., Mantri N. First Report of CRISPR/Cas9 Mediated DNA-Free Editing of 4CL and RVE7 Genes in Chickpea Protoplasts. Int. J. Mol. Sci. 2021;22:396. doi: 10.3390/ijms22010396. PubMed DOI PMC

Berger A., Guinand S., Boscari A., Puppo A., Brouquisse R. Medicago truncatula Phytoglobin 1.1 Controls Symbiotic Nodulation and Nitrogen Fixation via the Regulation of Nitric Oxide Concentration. New Phytol. 2020;227:84–98. doi: 10.1111/nph.16462. PubMed DOI PMC

Burén S., López-Torrejón G., Rubio L.M. Extreme Bioengineering to Meet the Nitrogen Challenge. Proc. Natl. Acad. Sci. USA. 2018;115:8849–8851. doi: 10.1073/pnas.1812247115. PubMed DOI PMC

Yang J., Xie X., Xiang N., Tian Z.-X., Dixon R., Wang Y.-P. Polyprotein Strategy for Stoichiometric Assembly of Nitrogen Fixation Components for Synthetic Biology. Proc. Natl. Acad. Sci. USA. 2018;115:E8509–E8517. doi: 10.1073/pnas.1804992115. PubMed DOI PMC

Battle A., Mostafavi S., Zhu X., Potash J.B., Weissman M.M., McCormick C., Haudenschild C.D., Beckman K.B., Shi J., Mei R., et al. Characterizing the Genetic Basis of Transcriptome Diversity through RNA-Sequencing of 922 Individuals. Genome Res. 2014;24:14–24. doi: 10.1101/gr.155192.113. PubMed DOI PMC

Sun Y., Xiao H. Identification of Alternative Splicing Events by RNA Sequencing in Early Growth Tomato Fruits. BMC Genom. 2015;16:948. doi: 10.1186/s12864-015-2128-6. PubMed DOI PMC

Deng T., Pang C., Lu X., Zhu P., Duan A., Tan Z., Huang J., Li H., Chen M., Liang X. De Novo Transcriptome Assembly of the Chinese Swamp Buffalo by RNA Sequencing and SSR Marker Discovery. PLoS ONE. 2016;11:e0147132. doi: 10.1371/journal.pone.0147132. PubMed DOI PMC

Zhang T., Huang L., Wang Y., Wang W., Zhao X., Zhang S., Zhang J., Hu F., Fu B., Li Z. Differential Transcriptome Profiling of Chilling Stress Response between Shoots and Rhizomes of Oryza Longistaminata Using RNA Sequencing. PLoS ONE. 2017;12:e0188625. doi: 10.1371/journal.pone.0188625. PubMed DOI PMC

Jin H., Dong D., Yang Q., Zhu D. Salt-Responsive Transcriptome Profiling of Suaeda Glauca via RNA Sequencing. PLoS ONE. 2016;11:e0150504. doi: 10.1371/journal.pone.0150504. PubMed DOI PMC

Lohani N., Singh M.B., Bhalla P.L. RNA-Seq Highlights Molecular Events Associated With Impaired Pollen-Pistil Interactions Following Short-Term Heat Stress in Brassica napus. Front. Plant Sci. 2021;11:622748. doi: 10.3389/fpls.2020.622748. PubMed DOI PMC

Zhang C., Li X., Wang Z., Zhang Z., Wu Z. Identifying Key Regulatory Genes of Maize Root Growth and Development by RNA Sequencing. Genomics. 2020;112:5157–5169. doi: 10.1016/j.ygeno.2020.09.030. PubMed DOI

Wang X., Liu C., Tu B., Li Y., Chen H., Zhang Q., Liu X. Characterization on a Novel Rolled Leaves and Short Petioles Soybean Mutant Based on Seq-BSA and RNA-Seq Analysis. J. Plant Biol. 2022;65:261–277. doi: 10.1007/s12374-020-09295-x. DOI

Bellés-Sancho P., Lardi M., Liu Y., Eberl L., Zamboni N., Bailly A., Pessi G. Metabolomics and Dual RNA-Sequencing on Root Nodules Revealed New Cellular Functions Controlled by Paraburkholderia phymatum NifA. Metabolites. 2021;11:455. doi: 10.3390/metabo11070455. PubMed DOI PMC

Cabeza R.A., Liese R., Lingner A., von Stieglitz I., Neumann J., Salinas-Riester G., Pommerenke C., Dittert K., Schulze J. RNA-Seq Transcriptome Profiling Reveals That Medicago truncatula Nodules Acclimate N 2 Fixation before Emerging P Deficiency Reaches the Nodules. J. Exp. Bot. 2014;65:6035–6048. doi: 10.1093/jxb/eru341. PubMed DOI PMC

Kusakin P.G., Serova T.A., Gogoleva N.E., Gogolev Y.V., Tsyganov V.E. Laser Microdissection of Pisum Sativum L. Nodules Followed by RNA-Seq Analysis Revealed Crucial Transcriptomic Changes during Infected Cell Differentiation. Agronomy. 2021;11:2504. doi: 10.3390/agronomy11122504. DOI

Yuan S.L., Li R., Chen H.F., Zhang C.J., Chen L.M., Hao Q.N., Chen S.L., Shan Z.H., Yang Z.L., Zhang X.J., et al. RNA-Seq Analysis of Nodule Development at Five Different Developmental Stages of Soybean (Glycine Max) Inoculated with Bradyrhizobium japonicum Strain 113-2. Sci. Rep. 2017;7:42248. doi: 10.1038/srep42248. PubMed DOI PMC

Ištvánek J., Jaroš M., Křenek A., Řepková J. Genome Assembly and Annotation for Red Clover (Trifolium pratense; Fabaceae) Am. J. Bot. 2014;101:327–337. doi: 10.3732/ajb.1300340. PubMed DOI

De Vega J.J., Ayling S., Hegarty M., Kudrna D., Goicoechea J.L., Ergon Å., Rognli O.A., Jones C., Swain M., Geurts R., et al. Red Clover (Trifolium pratense L.) Draft Genome Provides a Platform for Trait Improvement. Sci. Rep. 2015;5:17394. doi: 10.1038/srep17394. PubMed DOI PMC

Yates S.A., Swain M.T., Hegarty M.J., Chernukin I., Lowe M., Allison G.G., Ruttink T., Abberton M.T., Jenkins G., Skøt L. De Novo Assembly of Red Clover Transcriptome Based on RNA-Seq Data Provides Insight into Drought Response, Gene Discovery and Marker Identification. BMC Genom. 2014;15:453. doi: 10.1186/1471-2164-15-453. PubMed DOI PMC

Herbert D.B., Gross T., Rupp O., Becker A. Transcriptome Analysis Reveals Major Transcriptional Changes during Regrowth after Mowing of Red Clover (Trifolium pratense) BMC Plant Biol. 2021;21:95. doi: 10.1186/s12870-021-02867-0. PubMed DOI PMC

Chao Y., Xie L., Yuan J., Guo T., Li Y., Liu F., Han L. Transcriptome Analysis of Leaf Senescence in Red Clover (Trifolium pratense L.) Physiol. Mol. Biol. Plants. 2018;24:753–765. doi: 10.1007/s12298-018-0562-z. PubMed DOI PMC

Chao Y., Yuan J., Li S., Jia S., Han L., Xu L. Analysis of Transcripts and Splice Isoforms in Red Clover (Trifolium pratense L.) by Single-Molecule Long-Read Sequencing. BMC Plant Biol. 2018;18:300. doi: 10.1186/s12870-018-1534-8. PubMed DOI PMC

Shi K., Liu X., Pan X., Liu J., Gong W., Gong P., Cao M., Jia S., Wang Z. Unveiling the Complexity of Red Clover (Trifolium pratense L.) Transcriptome and Transcriptional Regulation of Isoflavonoid Biosynthesis Using Integrated Long- and Short-Read RNAseq. Int. J. Mol. Sci. 2021;22:12625. doi: 10.3390/ijms222312625. PubMed DOI PMC

Zhang J., Li J., Zou L., Li H. Transcriptome Analysis of Air Space-Type Variegation Formation in Trifolium Pratense. Int. J. Mol. Sci. 2022;23:7794. doi: 10.3390/ijms23147794. PubMed DOI PMC

Provorov N., Tikhonovich I. Genetic Resources for Improving Nitrogen Fixation in Legume-Rhizobia Symbiosis. Genet. Resour. Crop Evol. 2003;50:89–99. doi: 10.1023/A:1022957429160. DOI

Hardy R.W.F., Burns R.C., Holsten R.D. Applications of the Acetylene-Ethylene Assay for Measurement of Nitrogen Fixation. Soil Biol. Biochem. 1973;5:47–81. doi: 10.1016/0038-0717(73)90093-X. DOI

Unkovich M., Herridge D., Peoples M., Cadisch G., Boddey B., Giller K., Alves B., Chalk P. Measuring Plant-Associated Nitrogen Fixation in Agricultural Systems. Australian Centre for International Agricultural Research (ACIAR); Canberra, ACT, Australia: 2008.

Andrews S. FastQC: A Quality Control Tool for High Throughput Sequence Data. [(accessed on 29 October 2020)]. Available online: http://www.bioinformatics.babraham.ac.uk/projects/fastqc/

Bushnell B. BBMap: A Fast, Accurate, Splice-Aware Aligner. LBNL Report LBNL-7065E, Lawrence Berkeley National Laboratory. 2014. [(accessed on 2 November 2020)]. Available online: https://escholarship.org/uc/item/1h3515gn.

Dobin A., Davis C.A., Schlesinger F., Drenkow J., Zaleski C., Jha S., Batut P., Chaisson M., Gingeras T.R. STAR: Ultrafast Universal RNA-Seq Aligner. Bioinformatics. 2013;29:15–21. doi: 10.1093/bioinformatics/bts635. PubMed DOI PMC

Li H., Handsaker B., Wysoker A., Fennell T., Ruan J., Homer N., Marth G., Abecasis G., Durbin R. 1000 Genome Project Data Processing Subgroup The Sequence Alignment/Map Format and SAMtools. Bioinformatics. 2009;25:2078–2079. doi: 10.1093/bioinformatics/btp352. PubMed DOI PMC

Hartley S.W., Mullikin J.C. QoRTs: A Comprehensive Toolset for Quality Control and Data Processing of RNA-Seq Experiments. BMC Bioinform. 2015;16:224. doi: 10.1186/s12859-015-0670-5. PubMed DOI PMC

Trapnell C., Williams B.A., Pertea G., Mortazavi A., Kwan G., van Baren M.J., Salzberg S.L., Wold B.J., Pachter L. Transcript Assembly and Quantification by RNA-Seq Reveals Unannotated Transcripts and Isoform Switching during Cell Differentiation. Nat. Biotechnol. 2010;28:511–515. doi: 10.1038/nbt.1621. PubMed DOI PMC

Liao Y., Smyth G.K., Shi W. FeatureCounts: An Efficient General Purpose Program for Assigning Sequence Reads to Genomic Features. Bioinformatics. 2014;30:923–930. doi: 10.1093/bioinformatics/btt656. PubMed DOI

Love M.I., Huber W., Anders S. Moderated Estimation of Fold Change and Dispersion for RNA-Seq Data with DESeq2. Genome Biol. 2014;15:550. doi: 10.1186/s13059-014-0550-8. PubMed DOI PMC

Team R.S. RStudio, PBC; Boston, MA, USA: 2021. [(accessed on 10 November 2020)]. RStudio: Integrated Development for R. Available online: http://www.rstudio.com.

Goodstein D.M., Shu S., Howson R., Neupane R., Hayes R.D., Fazo J., Mitros T., Dirks W., Hellsten U., Putnam N., et al. Phytozome: A Comparative Platform for Green Plant Genomics. Nucleic Acids Res. 2012;40:D1178–D1186. doi: 10.1093/nar/gkr944. PubMed DOI PMC

Li J., Dai X., Liu T., Zhao P.X. LegumeIP: An Integrative Database for Comparative Genomics and Transcriptomics of Model Legumes. Nucleic Acids Res. 2012;40:D1221–D1229. doi: 10.1093/nar/gkr939. PubMed DOI PMC

Altschul S.F., Gish W., Miller W., Myers E.W., Lipman D.J. Basic Local Alignment Search Tool. J. Mol. Biol. 1990;215:403–410. doi: 10.1016/S0022-2836(05)80360-2. PubMed DOI

Boeckmann B. The SWISS-PROT Protein Knowledgebase and Its Supplement TrEMBL in 2003. Nucleic Acids Res. 2003;31:365–370. doi: 10.1093/nar/gkg095. PubMed DOI PMC

Kaur P., Bayer P.E., Milec Z., Vrána J., Yuan Y., Appels R., Edwards D., Batley J., Nichols P., Erskine W., et al. An Advanced Reference Genome of Trifolium Subterraneum L. Reveals Genes Related to Agronomic Performance. Plant Biotechnol. J. 2017;15:1034–1046. doi: 10.1111/pbi.12697. PubMed DOI PMC

Gotz S., Garcia-Gomez J.M., Terol J., Williams T.D., Nagaraj S.H., Nueda M.J., Robles M., Talon M., Dopazo J., Conesa A. High-Throughput Functional Annotation and Data Mining with the Blast2GO Suite. Nucleic Acids Res. 2008;36:3420–3435. doi: 10.1093/nar/gkn176. PubMed DOI PMC

Bioinformatics B., Valencia S. OmicsBox-Bioinformatics Made Easy. March. 2019;3:2019.

Koressaar T., Remm M. Enhancements and Modifications of Primer Design Program Primer3. Bioinformatics. 2007;23:1289–1291. doi: 10.1093/bioinformatics/btm091. PubMed DOI

Camacho C., Coulouris G., Avagyan V., Ma N., Papadopoulos J., Bealer K., Madden T.L. BLAST+: Architecture and Applications. BMC Bioinform. 2009;10:421. doi: 10.1186/1471-2105-10-421. PubMed DOI PMC

Maróti G., Downie J.A., Kondorosi É. Plant Cysteine-Rich Peptides That Inhibit Pathogen Growth and Control Rhizobial Differentiation in Legume Nodules. Curr. Opin. Plant Biol. 2015;26:57–63. doi: 10.1016/j.pbi.2015.05.031. PubMed DOI

Pruitt K.D., Tatusova T., Maglott D.R. NCBI Reference Sequences (RefSeq): A Curated Non-Redundant Sequence Database of Genomes, Transcripts and Proteins. Nucleic Acids Res. 2007;35:D61–D65. doi: 10.1093/nar/gkl842. PubMed DOI PMC

Lu S., Wang J., Chitsaz F., Derbyshire M.K., Geer R.C., Gonzales N.R., Gwadz M., Hurwitz D.I., Marchler G.H., Song J.S., et al. CDD/SPARCLE: The Conserved Domain Database in 2020. Nucleic Acids Res. 2020;48:D265–D268. doi: 10.1093/nar/gkz991. PubMed DOI PMC

Almagro Armenteros J.J., Tsirigos K.D., Sønderby C.K., Petersen T.N., Winther O., Brunak S., von Heijne G., Nielsen H. SignalP 5.0 Improves Signal Peptide Predictions Using Deep Neural Networks. Nat. Biotechnol. 2019;37:420–423. doi: 10.1038/s41587-019-0036-z. PubMed DOI

Almagro Armenteros J.J., Sønderby C.K., Sønderby S.K., Nielsen H., Winther O. DeepLoc: Prediction of Protein Subcellular Localization Using Deep Learning. Bioinformatics. 2017;33:3387–3395. doi: 10.1093/bioinformatics/btx431. PubMed DOI

Gasteiger E., Hoogland C., Gattiker A., Wilkins M.R., Appel R.D., Bairoch A. The Proteomics Protocols Handbook. Springer; Berlin/Heidelberg, Germany: 2005. Protein Identification and Analysis Tools on the ExPASy Server; pp. 571–607.

Kelley L.A., Mezulis S., Yates C.M., Wass M.N., Sternberg M.J.E. The Phyre2 Web Portal for Protein Modeling, Prediction and Analysis. Nat. Protoc. 2015;10:845–858. doi: 10.1038/nprot.2015.053. PubMed DOI PMC

Yang J., Anishchenko I., Park H., Peng Z., Ovchinnikov S., Baker D. Improved Protein Structure Prediction Using Predicted Interresidue Orientations. Proc. Natl. Acad. Sci. USA. 2020;117:1496–1503. doi: 10.1073/pnas.1914677117. PubMed DOI PMC

Qiao X., Li Q., Yin H., Qi K., Li L., Wang R., Zhang S., Paterson A.H. Gene Duplication and Evolution in Recurring Polyploidization–Diploidization Cycles in Plants. Genome Biol. 2019;20:38. doi: 10.1186/s13059-019-1650-2. PubMed DOI PMC

Langfelder P., Horvath S. WGCNA: An R Package for Weighted Correlation Network Analysis. BMC Bioinform. 2008;9:559. doi: 10.1186/1471-2105-9-559. PubMed DOI PMC

Shannon P. Cytoscape: A Software Environment for Integrated Models of Biomolecular Interaction Networks. Genome Res. 2003;13:2498–2504. doi: 10.1101/gr.1239303. PubMed DOI PMC

Maere S., Heymans K., Kuiper M. BiNGO: A Cytoscape Plugin to Assess Overrepresentation of Gene Ontology Categories in Biological Networks. Bioinformatics. 2005;21:3448–3449. doi: 10.1093/bioinformatics/bti551. PubMed DOI

Benjamini Y., Hochberg Y. Controlling the False Discovery Rate: A Practical and Powerful Approach to Multiple Testing. J. R. Stat. Soc. Ser. B Methodol. 1995;57:289–300. doi: 10.1111/j.2517-6161.1995.tb02031.x. DOI

Wang G., Oh D.-H., Dassanayake M. GOMCL: A Toolkit to Cluster, Evaluate, and Extract Non-Redundant Associations of Gene Ontology-Based Functions. BMC Bioinform. 2020;21:1–9. doi: 10.1186/s12859-020-3447-4. PubMed DOI PMC

Vance C.P. Legume Symbiotic Nitrogen Fixation: Agronomic Aspects. In: Spaink H.P., Kondorosi A., Hooykaas P.J.J., editors. The Rhizobiaceae. Springer Netherlands; Dordrecht, The Netherlands: 1998. pp. 509–530.

Sulieman S., Schulze J. The Efficiency of Nitrogen Fixation of the Model Legume Medicago truncatula (Jemalong A17) Is Low Compared to Medicago sativa. J. Plant Physiol. 2010;167:683–692. doi: 10.1016/j.jplph.2009.12.016. PubMed DOI

Yang Y., Zhao Q., Li X., Ai W., Liu D., Qi W., Zhang M., Yang C., Liao H. Characterization of Genetic Basis on Synergistic Interactions between Root Architecture and Biological Nitrogen Fixation in Soybean. Front. Plant Sci. 2017;8:1466. doi: 10.3389/fpls.2017.01466. PubMed DOI PMC

Smith G., Knight W., Peterson H. Variation among Inbred Lines of Crimson Clover for N2 Fixation (C2H2) Efficiency 1. Crop Sci. 1982;22:716–719. doi: 10.2135/cropsci1982.0011183X002200040005x. DOI

Luna R., Planchon C. Genotype x Bradyrhizobium japonicum Strain Interactions in Dinitrogen Fixation and Agronomic Traits of Soybean (Glycine max L. Merr.) Euphytica. 1995;86:127–134. doi: 10.1007/BF00022018. DOI

Peoples M.B., Baldock J.A. Nitrogen Dynamics of Pastures: Nitrogen Fixation Inputs, the Impact of Legumes on Soil Nitrogen Fertility, and the Contributions of Fixed Nitrogen to Australian Farming Systems. Aust. J. Exp. Agric. 2001;41:327. doi: 10.1071/EA99139. DOI

Yuan S., Li R., Chen S., Chen H., Zhang C., Chen L., Hao Q., Shan Z., Yang Z., Qiu D., et al. RNA-Seq Analysis of Differential Gene Expression Responding to Different Rhizobium Strains in Soybean (Glycine max) Roots. Front. Plant Sci. 2016;7:721. doi: 10.3389/fpls.2016.00721. PubMed DOI PMC

Cannon S.B., McKain M.R., Harkess A., Nelson M.N., Dash S., Deyholos M.K., Peng Y., Joyce B., Stewart C.N., Rolf M., et al. Multiple Polyploidy Events in the Early Radiation of Nodulating and Nonnodulating Legumes. Mol. Biol. Evol. 2015;32:193–210. doi: 10.1093/molbev/msu296. PubMed DOI PMC

Thilakarathna M.S., Papadopoulos Y.A., Grimmett M., Fillmore S.A.E., Crouse M., Prithiviraj B. Red Clover Varieties Show Nitrogen Fixing Advantage during the Early Stages of Seedling Development. Can. J. Plant Sci. 2018;98:517–526. doi: 10.1139/cjps-2017-0071. DOI

Soper F.M., Simon C., Jauss V. Measuring Nitrogen Fixation by the Acetylene Reduction Assay (ARA): Is 3 the Magic Ratio? Biogeochemistry. 2021;152:345–351. doi: 10.1007/s10533-021-00761-3. DOI

Roughley R., Dart P. Reduction of Acetylene by Nodules of Trifolium Subterraneum as Affected by Root Temperature, Rhizobium Strain and Host Cultivar. Arch. Für Mikrobiol. 1969;69:171–179. doi: 10.1007/BF00409761. DOI

Bergersen F.J. The Quantitative Relationship between Nitrogen Fixation and the Acetylene-Reduction Assay. Aust. J. Biol. Sci. 1970;23:1015–1026. doi: 10.1071/BI9701015. DOI

Anderson M.D., Ruess R.W., Uliassi D.D., Mitchell J.S. Estimating N2 Fixation in Two Species of Alnus in Interior Alaska Using Acetylene Reduction and 15N2 Uptake. Ecoscience. 2004;11:102–112. doi: 10.1080/11956860.2004.11682814. DOI

Ayanaba A., Lawson T. Diurnal Changes in Acetylene Reduction in Field-Grown Cowpeas and Soybeans. Soil Biol. Biochem. 1977;9:125–129. doi: 10.1016/0038-0717(77)90048-7. DOI

Zapata F., Danso F., Hardarson G., Fried M. Nitrogen Fixation and Translocation in Field-Grown Fababean. Agron. J. 1987;79:505–509. doi: 10.2134/agronj1987.00021962007900030020x. DOI

Vessey J.K. Measurement of Nitrogenase Activity in Legume Root Nodules: In Defense of the Acetylene Reduction Assay. Plant Soil. 1994;158:151–162. doi: 10.1007/BF00009490. DOI

Young N.D., Cannon S.B., Sato S., Kim D., Cook D.R., Town C.D., Roe B.A., Tabata S. Sequencing the Genespaces of Medicago truncatula and Lotus japonicus. Plant Physiol. 2005;137:1174–1181. doi: 10.1104/pp.104.057034. PubMed DOI PMC

Mumm R., Posthumus M.A., Dicke M. Significance of Terpenoids in Induced Indirect Plant Defence against Herbivorous Arthropods. Plant Cell Environ. 2008;31:575–585. doi: 10.1111/j.1365-3040.2008.01783.x. PubMed DOI

Umehara M., Hanada A., Yoshida S., Akiyama K., Arite T., Takeda-Kamiya N., Magome H., Kamiya Y., Shirasu K., Yoneyama K., et al. Inhibition of Shoot Branching by New Terpenoid Plant Hormones. Nature. 2008;455:195–200. doi: 10.1038/nature07272. PubMed DOI

Zhou Y., Stuart-Williams H., Grice K., Kayler Z.E., Zavadlav S., Vogts A., Rommerskirchen F., Farquhar G.D., Gessler A. Allocate Carbon for a Reason: Priorities Are Reflected in the 13C/12C Ratios of Plant Lipids Synthesized via Three Independent Biosynthetic Pathways. Phytochemistry. 2015;111:14–20. doi: 10.1016/j.phytochem.2014.12.005. PubMed DOI

Ali M., Miao L., Hou Q., Darwish D.B., Alrdahe S.S., Ali A., Benedito V.A., Tadege M., Wang X., Zhao J. Overexpression of Terpenoid Biosynthesis Genes From Garden Sage (Salvia officinalis) Modulates Rhizobia Interaction and Nodulation in Soybean. Front. Plant Sci. 2021;12:783269. doi: 10.3389/fpls.2021.783269. PubMed DOI PMC

Foo E., Davies N.W. Strigolactones Promote Nodulation in Pea. Planta. 2011;234:1073–1081. doi: 10.1007/s00425-011-1516-7. PubMed DOI

Foo E., Yoneyama K., Hugill C.J., Quittenden L.J., Reid J.B. Strigolactones and the Regulation of Pea Symbioses in Response to Nitrate and Phosphate Deficiency. Mol. Plant. 2013;6:76–87. doi: 10.1093/mp/sss115. PubMed DOI

ur Rehman N., Ali M., Ahmad M.Z., Liang G., Zhao J. Strigolactones Promote Rhizobia Interaction and Increase Nodulation in Soybean (Glycine max) Microb. Pathog. 2018;114:420–430. doi: 10.1016/j.micpath.2017.11.049. PubMed DOI

Akbar A., Han B., Khan A.H., Feng C., Ullah A., Khan A.S., He L., Yang X. A Transcriptomic Study Reveals Salt Stress Alleviation in Cotton Plants upon Salt Tolerant PGPR Inoculation. Environ. Exp. Bot. 2022;200:104928. doi: 10.1016/j.envexpbot.2022.104928. DOI

Hubbard C.J., Li B., McMinn R., Brock M.T., Maignien L., Ewers B.E., Kliebenstein D., Weinig C. The Effect of Rhizosphere Microbes Outweighs Host Plant Genetics in Reducing Insect Herbivory. Mol. Ecol. 2019;28:1801–1811. doi: 10.1111/mec.14989. PubMed DOI

Seifert G.J. Nucleotide Sugar Interconversions and Cell Wall Biosynthesis: How to Bring the inside to the Outside. Curr. Opin. Plant Biol. 2004;7:277–284. doi: 10.1016/j.pbi.2004.03.004. PubMed DOI

O’Donoghue E.M., Somerfield S.D., Watson L.M., Brummell D.A., Hunter D.A. Galactose Metabolism in Cell Walls of Opening and Senescing Petunia Petals. Planta. 2009;229:709–721. doi: 10.1007/s00425-008-0862-6. PubMed DOI

Lee D.-K., Ahn S., Cho H.Y., Yun H.Y., Park J.H., Lim J., Lee J., Kwon S.W. Metabolic Response Induced by Parasitic Plant-Fungus Interactions Hinder Amino Sugar and Nucleotide Sugar Metabolism in the Host. Sci. Rep. 2016;6:37434. doi: 10.1038/srep37434. PubMed DOI PMC

La Camera S., Gouzerh G., Dhondt S., Hoffmann L., Fritig B., Legrand M., Heitz T. Metabolic Reprogramming in Plant Innate Immunity: The Contributions of Phenylpropanoid and Oxylipin Pathways. Immunol. Rev. 2004;198:267–284. doi: 10.1111/j.0105-2896.2004.0129.x. PubMed DOI

Bais H.P., Vepachedu R., Gilroy S., Callaway R.M., Vivanco J.M. Allelopathy and Exotic Plant Invasion: From Molecules and Genes to Species Interactions. Science. 2003;301:1377–1380. doi: 10.1126/science.1083245. PubMed DOI

Vogt T. Phenylpropanoid Biosynthesis. Mol. Plant. 2010;3:2–20. doi: 10.1093/mp/ssp106. PubMed DOI

Peck M.C., Fisher R.F., Long S.R. Diverse Flavonoids Stimulate NodD1 Binding to Nod Gene Promoters in Sinorhizobium meliloti. J. Bacteriol. 2006;188:5417–5427. doi: 10.1128/JB.00376-06. PubMed DOI PMC

Wasson A.P., Pellerone F.I., Mathesius U. Silencing the Flavonoid Pathway in Medicago truncatula Inhibits Root Nodule Formation and Prevents Auxin Transport Regulation by Rhizobia. Plant Cell. 2006;18:1617–1629. doi: 10.1105/tpc.105.038232. PubMed DOI PMC

Mithöfer A. Suppression of Plant Defence in Rhizobia–Legume Symbiosis. Trends Plant Sci. 2002;7:440–444. doi: 10.1016/S1360-1385(02)02336-1. PubMed DOI

Staehelin C., Krishnan H.B. Nodulation Outer Proteins: Double-Edged Swords of Symbiotic Rhizobia. Biochem. J. 2015;470:263–274. doi: 10.1042/BJ20150518. PubMed DOI

Avenhaus U., Cabeza R.A., Liese R., Lingner A., Dittert K., Salinas-Riester G., Pommerenke C., Schulze J. Short-Term Molecular Acclimation Processes of Legume Nodules to Increased External Oxygen Concentration. Front. Plant Sci. 2016;6:1133. doi: 10.3389/fpls.2015.01133. PubMed DOI PMC

Milligan A.J., Berman-Frank I., Gerchman Y., Dismukes G.C., Falkowski P.G. Light-Dependent Oxygen Consumption in Nitrogen-Fixing Cyanobacteria Plays a Key Role in Nitrogenase Protection. J. Phycol. 2007;43:845–852. doi: 10.1111/j.1529-8817.2007.00395.x. DOI

Mylona P., Pawlowski K., Bisseling’ T. Symbiotic Nitrogen Fixation. Plant Cell. 1995;7:869–885. doi: 10.2307/3870043. PubMed DOI PMC

Young N.D., Debellé F., Oldroyd G.E.D., Geurts R., Cannon S.B., Udvardi M.K., Benedito V.A., Mayer K.F.X., Gouzy J., Schoof H., et al. The Medicago Genome Provides Insight into the Evolution of Rhizobial Symbioses. Nature. 2011;480:520–524. doi: 10.1038/nature10625. PubMed DOI PMC

Mandal D., Sinharoy S. A Toolbox for Nodule Development Studies in Chickpea: A Hairy-Root Transformation Protocol and an Efficient Laboratory Strain of Mesorhizobium Sp. Mol. Plant-Microbe Interact. 2019;32:367–378. doi: 10.1094/MPMI-09-18-0264-TA. PubMed DOI

Panchy N., Lehti-Shiu M., Shiu S.-H. Evolution of Gene Duplication in Plants. Plant Physiol. 2016;171:2294–2316. doi: 10.1104/pp.16.00523. PubMed DOI PMC

Li Q.-G., Zhang L., Li C., Dunwell J.M., Zhang Y.-M. Comparative Genomics Suggests That an Ancestral Polyploidy Event Leads to Enhanced Root Nodule Symbiosis in the Papilionoideae. Mol. Biol. Evol. 2013;30:2602–2611. doi: 10.1093/molbev/mst152. PubMed DOI

Hanada K., Zou C., Lehti-Shiu M.D., Shinozaki K., Shiu S.-H. Importance of Lineage-Specific Expansion of Plant Tandem Duplicates in the Adaptive Response to Environmental Stimuli. Plant Physiol. 2008;148:993–1003. doi: 10.1104/pp.108.122457. PubMed DOI PMC

Wang Y., Ficklin S.P., Wang X., Feltus F.A., Paterson A.H. Large-Scale Gene Relocations Following an Ancient Genome Triplication Associated with the Diversification of Core Eudicots. PLoS ONE. 2016;11:e0155637. doi: 10.1371/journal.pone.0155637. PubMed DOI PMC

Fuller T., Langfelder P., Presson A., Horvath S. Review of Weighted Gene Coexpression Network Analysis. In: Lu H.H.-S., Schölkopf B., Zhao H., editors. Handbook of Statistical Bioinformatics. Springer Berlin Heidelberg; Berlin/Heidelberg, Germany: 2011. pp. 369–388.

Battenberg K., Potter D., Tabuloc C.A., Chiu J.C., Berry A.M. Comparative Transcriptomic Analysis of Two Actinorhizal Plants and the Legume Medicago truncatula Supports the Homology of Root Nodule Symbioses and Is Congruent With a Two-Step Process of Evolution in the Nitrogen-Fixing Clade of Angiosperms. Front. Plant Sci. 2018;9:1256. doi: 10.3389/fpls.2018.01256. PubMed DOI PMC

Soltis D.E., Soltis P.S., Morgan D.R., Swensen S.M., Mullin B.C., Dowd J.M., Martin P.G. Chloroplast Gene Sequence Data Suggest a Single Origin of the Predisposition for Symbiotic Nitrogen Fixation in Angiosperms. Proc. Natl. Acad. Sci. USA. 1995;92:2647–2651. doi: 10.1073/pnas.92.7.2647. PubMed DOI PMC

Fedorova M., van de Mortel J., Matsumoto P.A., Cho J., Town C.D., VandenBosch K.A., Gantt J.S., Vance C.P. Genome-Wide Identification of Nodule-Specific Transcripts in the Model Legume Medicago truncatula. Plant Physiol. 2002;130:519–537. doi: 10.1104/pp.006833. PubMed DOI PMC

El Yahyaoui F., Küster H., Ben Amor B., Hohnjec N., Pühler A., Becker A., Gouzy J., Vernié T., Gough C., Niebel A., et al. Expression Profiling in Medicago truncatula Identifies More Than 750 Genes Differentially Expressed during Nodulation, Including Many Potential Regulators of the Symbiotic Program. Plant Physiol. 2004;136:3159–3176. doi: 10.1104/pp.104.043612. PubMed DOI PMC

Lee H., Hur C.-G., Oh C.J., Kim H.B., Park S.-Y., An C.S. Analysis of the Root Nodule-Enhanced Transcriptome in Soybean. Mol. Cells. 2004;18:53–62. PubMed

Liu J., Rasing M., Zeng T., Klein J., Kulikova O., Bisseling T. NIN Is Essential for Development of Symbiosomes, Suppression of Defence and Premature Senescence in Medicago truncatula Nodules. New Phytol. 2021;230:290–303. doi: 10.1111/nph.17215. PubMed DOI PMC

Benedito V.A., Torres-Jerez I., Murray J.D., Andriankaja A., Allen S., Kakar K., Wandrey M., Verdier J., Zuber H., Ott T., et al. A Gene Expression Atlas of the Model Legume Medicago truncatula. Plant J. 2008;55:504–513. doi: 10.1111/j.1365-313X.2008.03519.x. PubMed DOI

Høgslund N., Radutoiu S., Krusell L., Voroshilova V., Hannah M.A., Goffard N., Sanchez D.H., Lippold F., Ott T., Sato S., et al. Dissection of Symbiosis and Organ Development by Integrated Transcriptome Analysis of Lotus japonicus Mutant and Wild-Type Plants. PLoS ONE. 2009;4:e6556. doi: 10.1371/journal.pone.0006556. PubMed DOI PMC

Libault M., Farmer A., Joshi T., Takahashi K., Langley R.J., Franklin L.D., He J., Xu D., May G., Stacey G. An Integrated Transcriptome Atlas of the Crop Model Glycine max, and Its Use in Comparative Analyses in Plants: Soybean Transcriptome Atlas. Plant J. 2010;63:86–99. doi: 10.1111/j.1365-313X.2010.04222.x. PubMed DOI

Maunoury N., Redondo-Nieto M., Bourcy M., Van de Velde W., Alunni B., Laporte P., Durand P., Agier N., Marisa L., Vaubert D., et al. Differentiation of Symbiotic Cells and Endosymbionts in Medicago truncatula Nodulation Are Coupled to Two Transcriptome-Switches. PLoS ONE. 2010;5:e9519. doi: 10.1371/journal.pone.0009519. PubMed DOI PMC

Breakspear A., Liu C., Roy S., Stacey N., Rogers C., Trick M., Morieri G., Mysore K.S., Wen J., Oldroyd G.E.D., et al. The Root Hair “Infectome” of Medicago truncatula Uncovers Changes in Cell Cycle Genes and Reveals a Requirement for Auxin Signaling in Rhizobial Infection. Plant Cell. 2014;26:4680–4701. doi: 10.1105/tpc.114.133496. PubMed DOI PMC

Larrainzar E., Riely B.K., Kim S.C., Carrasquilla-Garcia N., Yu H.-J., Hwang H.-J., Oh M., Kim G.B., Surendrarao A.K., Chasman D., et al. Deep Sequencing of the Medicago truncatula Root Transcriptome Reveals a Massive and Early Interaction between Nodulation Factor and Ethylene Signals. Plant Physiol. 2015;169:233–265. doi: 10.1104/pp.15.00350. PubMed DOI PMC

Damiani I., Drain A., Guichard M., Balzergue S., Boscari A., Boyer J.-C., Brunaud V., Cottaz S., Rancurel C., Da Rocha M., et al. Nod Factor Effects on Root Hair-Specific Transcriptome of Medicago truncatula: Focus on Plasma Membrane Transport Systems and Reactive Oxygen Species Networks. Front. Plant Sci. 2016;7:794. doi: 10.3389/fpls.2016.00794. PubMed DOI PMC

Jardinaud M.-F., Boivin S., Rodde N., Catrice O., Kisiala A., Lepage A., Moreau S., Roux B., Cottret L., Sallet E., et al. A Laser Dissection-RNAseq Analysis Highlights the Activation of Cytokinin Pathways by Nod Factors in the Medicago truncatula Root Epidermis. Plant Physiol. 2016;171:2256–2276. doi: 10.1104/pp.16.00711. PubMed DOI PMC

Schiessl K., Lilley J.L.S., Lee T., Tamvakis I., Kohlen W., Bailey P.C., Thomas A., Luptak J., Ramakrishnan K., Carpenter M.D., et al. NODULE INCEPTION Recruits the Lateral Root Developmental Program for Symbiotic Nodule Organogenesis in Medicago truncatula. Curr. Biol. 2019;29:3657–3668. doi: 10.1016/j.cub.2019.09.005. PubMed DOI PMC

Rudaya E.S., Kozyulina P.Y., Pavlova O.A., Dolgikh A.V., Ivanova A.N., Dolgikh E.A. Regulation of the Later Stages of Nodulation Stimulated by IPD3/CYCLOPS Transcription Factor and Cytokinin in Pea Pisum sativum L. Plants. 2021;11:56. doi: 10.3390/plants11010056. PubMed DOI PMC