Viroids are small non-capsidated, single-stranded, covalently-closed circular noncoding RNA replicons of 239-401 nucleotides that exploit host factors for their replication, and some cause disease in several economically important crop plants, while others appear to be benign. The proposed mechanisms of viroid pathogenesis include direct interaction of the genomic viroid RNA with host factors and post-transcriptional or transcriptional gene silencing via viroid-derived small RNAs (vd-sRNAs) generated by the host defensive machinery. Humulus lupulus (hop) plants are hosts to several viroids among which Hop latent viroid (HLVd) and Citrus bark cracking viroid (CBCVd) are attractive model systems for the study of viroid-host interactions due to the symptomless infection of the former and severe symptoms induced by the latter in this indicator host. To better understand their interactions with hop plant, a comparative transcriptomic analysis based on RNA sequencing (RNA-seq) was performed to reveal the transcriptional alterations induced as a result of single HLVd and CBCVd infection in hop. Additionally, the effect of HLVd on the aggressiveness of CBCVd that underlies severe stunting in hop in a mixed infection was studied by transcriptomic analysis. Our analysis revealed that CBCVd infection resulted in dynamic changes in the activity of genes as compared to single HLVd infection and their mixed infection. The differentially expressed genes that are involved in defense, phytohormone signaling, photosynthesis and chloroplasts, RNA regulation, processing and binding; protein metabolism and modification; and other mechanisms were more modulated in the CBCVd infection of hop. Nevertheless, Gene Ontology (GO) classification and pathway enrichment analysis showed that the expression of genes involved in the proteolysis mechanism is more active in a mixed infection as compared to a single one, suggesting co-infecting viroids may result in interference with host factors more prominently. Collectively, our results provide a deep transcriptome of hop and insight into complex single HLVd, CBCVd, and their coinfection in hop-plant interactions.

Biology Centre of the Czech Academy of Sciences Institute of Plant Molecular Biology Department of Molecular Genetics Branišovská 31 37005 České Budějovice Czech Republic

Slovenian Institute of Hop Research and Brewing Plant Protection Department Cesta Žalskega tabora 2 SI 3310 Žalec Slovenia

University of Ljubljana Biotechnical Faculty Department of Agronomy Jamnikarjeva 101 SI 1000 Ljubljana Slovenia

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Flores R., Minoia S., Carbonell A., Gisel A., Delgado S., López-Carrasco A., Navarro B., Di Serio F. Viroids, the simplest RNA replicons: How they manipulate their hosts for being propagated and how their hosts react for containing the infection. Virus Res. 2015;209:136–145. doi: 10.1016/j.virusres.2015.02.027. PubMed DOI

Gago-Zachert S. Viroids, infectious long non-coding RNAs with autonomous replication. Virus Res. 2016;212:12–24. doi: 10.1016/j.virusres.2015.08.018. PubMed DOI

Gago S., Elena S.F., Flores R., Sanjuán R. Extremely High Mutation Rate of a Hammerhead Viroid. Science. 2009;323:1308. doi: 10.1126/science.1169202. PubMed DOI

Kovalskaya N., Hammond R.W. Molecular biology of viroid–host interactions and disease control strategies. Plant Sci. 2014;228:48–60. doi: 10.1016/j.plantsci.2014.05.006. PubMed DOI

Flores R., Hernández C., de Alba A.E.M., Daròs J.-A., Serio F. Di Viroids and Viroid-Host Interactions. Annu. Rev. Phytopathol. 2005;43:117–139. doi: 10.1146/annurev.phyto.43.040204.140243. PubMed DOI

Schindler I.-M., Mühlbach H.-P. Involvement of nuclear DNA-dependent RNA polymerases in potato spindle tuber viroid replication: A reevaluation. Plant Sci. 1992;84:221–229. doi: 10.1016/0168-9452(92)90138-C. DOI

Owens R.A., Diener T.O. RNA intermediates in potato spindle tuber viroid replication. Proc. Natl. Acad. Sci. USA. 1982;79:113–117. doi: 10.1073/pnas.79.1.113. PubMed DOI PMC

Daròs J.A., Marcos J.F., Hernández C., Flores R. Replication of avocado sunblotch viroid: Evidence for a symmetric pathway with two rolling circles and hammerhead ribozyme processing. Proc. Natl. Acad. Sci. USA. 1994;91:12813–12817. doi: 10.1073/pnas.91.26.12813. PubMed DOI PMC

Delgado S., Martínez de Alba A.E., Hernández C., Flores R. A short double-stranded RNA motif of Peach latent mosaic viroid contains the initiation and the self-cleavage sites of both polarity strands. J. Virol. 2005;79:12934–12943. doi: 10.1128/JVI.79.20.12934-12943.2005. PubMed DOI PMC

Wassenegger M., Heimes S., Riedel L., Sänger H.L. RNA-directed de novo methylation of genomic sequences in plants. Cell. 1994;76:567–576. doi: 10.1016/0092-8674(94)90119-8. PubMed DOI

Gómez G., Martínez G., Pallás V. Interplay between viroid-induced pathogenesis and RNA silencing pathways. Trends Plant Sci. 2009;14:264–269. doi: 10.1016/j.tplants.2009.03.002. PubMed DOI

Matoušek J., Siglová K., Jakše J., Radišek S., Brass J.R.J., Tsushima T., Guček T., Duraisamy G.S., Sano T., Steger G. Propagation and some physiological effects of Citrus bark cracking viroid and Apple fruit crinkle viroid in multiple infected hop (Humulus lupulus L.) J. Plant Physiol. 2017;213:166–177. doi: 10.1016/j.jplph.2017.02.014. PubMed DOI

Tomassoli L., Luison D., Luigi M., Costantini E., Mangiaracina P., Faggioli F. Acta Horticulturae. International Society for Horticultural Science (ISHS); Leuven, Belgium: 2015. Supportive and antagonistic interactions among pospiviroids infecting solanaceous ornamentals; pp. 71–77.

Lin C.-Y., Wu M.-L., Shen T.-L., Hung T.-H. A mutual titer-enhancing relationship and similar localization patterns between Citrus exocortis viroid and Hop stunt viroid co-infecting two citrus cultivars. Virol. J. 2015;12:142. doi: 10.1186/s12985-015-0357-6. PubMed DOI PMC

Karabín M., Hudcová T., Jelínek L., Dostálek P. Biologically Active Compounds from Hops and Prospects for Their Use. Compr. Rev. Food Sci. Food Saf. 2016;15:542–567. doi: 10.1111/1541-4337.12201. PubMed DOI

Sano T., Mimura R., Ohshima K. Phylogenetic Analysis of Hop and Grapevine Isolates of Hop Stunt Viroid Supports a Grapevine Origin for Hop Stunt Disease. Virus Genes. 2001;22:53–59. doi: 10.1023/A:1008182302704. PubMed DOI

Sano T., Isono S., Matsuki K., Kawaguchi-Ito Y., Tanaka K., Kondo K., Iijima A., Bar-Joseph M. Vegetative propagation and its possible role as a genetic bottleneck in the shaping of the apple fruit crinkle viroid populations in apple and hop plants. Virus Genes. 2008;37:298–303. doi: 10.1007/s11262-008-0270-9. PubMed DOI

Puchta H., Ramm K., Sänger H.L. The molecular structure of hop latent viroid (HLV), a new viroid occurring worldwide in hops. Nucleic Acids Res. 1988;16:4197–4216. doi: 10.1093/nar/16.10.4197. PubMed DOI PMC

Jakse J., Radisek S., Pokorn T., Matousek J., Javornik B. Deep-sequencing revealed Citrus bark cracking viroid (CBCVd) as a highly aggressive pathogen on hop. Plant Pathol. 2015;64:831–842. doi: 10.1111/ppa.12325. DOI

Barbara D.J., Morton A., Adams A.N., Green C.P. Some effects of hop latent viroid on two cultivars of hop (Humulus lupulus) in the UK. Ann. Appl. Biol. 1990;117:359–366. doi: 10.1111/j.1744-7348.1990.tb04222.x. DOI

Sano T., Yoshida H., Goshono M., Monma T., Kawasaki H., Ishizaki K. Characterization of a new viroid strain from hops: Evidence for viroid speciation by isolation in different host species. J. Gen. Plant Pathol. 2004;70:181–187. doi: 10.1007/s10327-004-0105-z. DOI

Kappagantu M., Nelson M.E., Bullock J.M., Kenny S.T., Eastwell K.C. Hop Stunt Viroid: Effects on Vegetative Growth and Yield of Hop Cultivars, and Its Distribution in Central Washington State. Plant Dis. 2017;101:607–612. doi: 10.1094/PDIS-06-16-0884-RE. PubMed DOI

Mishra A.K., Duraisamy G.S., Matoušek J., Radisek S., Javornik B., Jakse J. Identification and characterization of microRNAs in Humulus lupulus using high-throughput sequencing and their response to Citrus bark cracking viroid (CBCVd) infection. BMC Genom. 2016;17:919. doi: 10.1186/s12864-016-3271-4. PubMed DOI PMC

Itaya A., Matsuda Y., Gonzales R.A., Nelson R.S., Ding B. Potato spindle tuber viroid Strains of Different Pathogenicity Induces and Suppresses Expression of Common and Unique Genes in Infected Tomato. Mol. Plant-Microbe Interact. 2002;15:990–999. doi: 10.1094/MPMI.2002.15.10.990. PubMed DOI

Owens R.A., Tech K.B., Shao J.Y., Sano T., Baker C.J. Global Analysis of Tomato Gene Expression During Potato spindle tuber viroid Infection Reveals a Complex Array of Changes Affecting Hormone Signaling. Mol. Plant-Microbe Interact. 2012;25:582–598. doi: 10.1094/MPMI-09-11-0258. PubMed DOI

Więsyk A., Iwanicka-Nowicka R., Fogtman A., Zagórski-Ostoja W., Góra-Sochacka A. Time-Course Microarray Analysis Reveals Differences between Transcriptional Changes in Tomato Leaves Triggered by Mild and Severe Variants of Potato Spindle Tuber Viroid. Viruses. 2018;10:257. doi: 10.3390/v10050257. PubMed DOI PMC

Pokorn T., Radišek S., Javornik B., Štajner N., Jakše J. Development of hop transcriptome to support research into host-viroid interactions. PLoS ONE. 2017;12:e0184528. doi: 10.1371/journal.pone.0184528. PubMed DOI PMC

Zhao S., Fung-Leung W.-P., Bittner A., Ngo K., Liu X. Comparison of RNA-Seq and microarray in transcriptome profiling of activated T cells. PLoS ONE. 2014;9:e78644. doi: 10.1371/journal.pone.0078644. PubMed DOI PMC

Katsarou K., Wu Y., Zhang R., Bonar N., Morris J., Hedley P.E., Bryan G.J., Kalantidis K., Hornyik C. Insight on Genes Affecting Tuber Development in Potato upon Potato spindle tuber viroid (PSTVd) Infection. PLoS ONE. 2016;11:e0150711. doi: 10.1371/journal.pone.0150711. PubMed DOI PMC

Zheng Y., Wang Y., Ding B., Fei Z. Comprehensive Transcriptome Analyses Reveal that Potato Spindle Tuber Viroid Triggers Genome-Wide Changes in Alternative Splicing, Inducible trans-Acting Activity of Phased Secondary Small Interfering RNAs, and Immune Responses. J. Virol. 2017;91:e00247-17. doi: 10.1128/JVI.00247-17. PubMed DOI PMC

Herranz M.C., Niehl A., Rosales M., Fiore N., Zamorano A., Granell A., Pallas V. A remarkable synergistic effect at the transcriptomic level in peach fruits doubly infected by prunus necrotic ringspot virus and peach latent mosaic viroid. Virol. J. 2013;10:164. doi: 10.1186/1743-422X-10-164. PubMed DOI PMC

Mishra K.A., Kumar A., Mishra D., Nath S.V., Jakše J., Kocábek T., Killi K.U., Morina F., Matoušek J. Genome-Wide Transcriptomic Analysis Reveals Insights into the Response to Citrus bark cracking viroid (CBCVd) in Hop (Humulus lupulus L.) Viruses. 2018;10:570. doi: 10.3390/v10100570. PubMed DOI PMC

Kappagantu M., Bullock J.M., Nelson M.E., Eastwell K.C. Hop stunt viroid: Effect on Host (Humulus lupulus) Transcriptome and Its Interactions with Hop Powdery Mildew (Podospheara macularis) Mol. Plant-Microbe Interact. 2017;30:842–851. doi: 10.1094/MPMI-03-17-0071-R. PubMed DOI

Eastwell K.C., Nelson M.E. Occurrence of Viroids in Commercial Hop (Humulus lupulus L.) Production Areas of Washington State. Plant Health Prog. 2007;8:1. doi: 10.1094/PHP-2007-1127-01-RS. DOI

Bedre R., Mangu V.R., Srivastava S., Sanchez L.E., Baisakh N. Transcriptome analysis of smooth cordgrass (Spartina alterniflora Loisel), a monocot halophyte, reveals candidate genes involved in its adaptation to salinity. BMC Genom. 2016;17:657. doi: 10.1186/s12864-016-3017-3. PubMed DOI PMC

Garg R., Patel R.K., Tyagi A.K., Jain M. De novo assembly of chickpea transcriptome using short reads for gene discovery and marker identification. DNA Res. 2011;18:53–63. doi: 10.1093/dnares/dsq028. PubMed DOI PMC

Tao X., Gu Y.-H., Wang H.-Y., Zheng W., Li X., Zhao C.-W., Zhang Y.-Z. Digital Gene Expression Analysis Based on Integrated De Novo Transcriptome Assembly of Sweet Potato [Ipomoea batatas (L.) Lam.] PLoS ONE. 2012;7:e36234. doi: 10.1371/journal.pone.0036234. PubMed DOI PMC

Tranchida-Lombardo V., Aiese Cigliano R., Anzar I., Landi S., Palombieri S., Colantuono C., Bostan H., Termolino P., Aversano R., Batelli G., et al. Whole-genome re-sequencing of two Italian tomato landraces reveals sequence variations in genes associated with stress tolerance, fruit quality and long shelf-life traits. DNA Res. 2018;25:149–160. doi: 10.1093/dnares/dsx045. PubMed DOI PMC

Simão F.A., Waterhouse R.M., Ioannidis P., Kriventseva E.V., Zdobnov E.M. BUSCO: Assessing genome assembly and annotation completeness with single-copy orthologs. Bioinformatics. 2015;31:3210–3212. doi: 10.1093/bioinformatics/btv351. PubMed DOI

Mishra A.K., Duraisamy G.S., Khare M., Kocábek T., Jakse J., Bříza J., Patzak J., Sano T., Matoušek J. Genome-wide transcriptome profiling of transgenic hop (Humulus lupulus L.) constitutively overexpressing HlWRKY1 and HlWDR1 transcription factors. BMC Genom. 2018;19:739. doi: 10.1186/s12864-018-5125-8. PubMed DOI PMC

Lincoln J.E., Sanchez J.P., Zumstein K., Gilchrist D.G. Plant and animal PR1 family members inhibit programmed cell death and suppress bacterial pathogens in plant tissues. Mol. Plant Pathol. 2018;19:2111–2123. doi: 10.1111/mpp.12685. PubMed DOI PMC

Zhong R., Kays S.J., Schroeder B.P., Ye Z.-H. Mutation of a chitinase-like gene causes ectopic deposition of lignin, aberrant cell shapes, and overproduction of ethylene. Plant Cell. 2002;14:165–179. doi: 10.1105/tpc.010278. PubMed DOI PMC

Hermans C., Porco S., Verbruggen N., Bush D.R. Chitinase-Like Protein CTL1 Plays a Role in Altering Root System Architecture in Response to Multiple Environmental Conditions. Plant Physiol. 2010;152:904–917. doi: 10.1104/pp.109.149849. PubMed DOI PMC

Grabherr M.G., Haas B.J., Yassour M., Levin J.Z., Thompson D.A., Amit I., Adiconis X., Fan L., Raychowdhury R., Zeng Q., et al. Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nat. Biotechnol. 2011;29:644. doi: 10.1038/nbt.1883. PubMed DOI PMC

Robinson M.D., Oshlack A., Young M.D. From RNA-seq reads to differential expression results. Genome Biol. 2010;11:220. doi: 10.1186/gb-2010-11-12-220. PubMed DOI PMC

Macho A.P., Zipfel C. Plant PRRs and the Activation of Innate Immune Signaling. Mol. Cell. 2014;54:263–272. doi: 10.1016/j.molcel.2014.03.028. PubMed DOI

Zvereva A.S., Pooggin M.M. Silencing and innate immunity in plant defense against viral and non-viral pathogens. Viruses. 2012;4:2578–2597. doi: 10.3390/v4112578. PubMed DOI PMC

Gao X., Cox K.L., Jr., He P. Functions of Calcium-Dependent Protein Kinases in Plant Innate Immunity. Plants. 2014;3:160–176. doi: 10.3390/plants3010160. PubMed DOI PMC

Zeng H., Xu L., Singh A., Wang H., Du L., Poovaiah B.W. Involvement of calmodulin and calmodulin-like proteins in plant responses to abiotic stresses. Front. Plant Sci. 2015;6:600. doi: 10.3389/fpls.2015.00600. PubMed DOI PMC

Krug R.M. Viral proteins that bind double-stranded RNA: Countermeasures against host antiviral responses. J. Interferon Cytokine Res. 2014;34:464–468. doi: 10.1089/jir.2014.0005. PubMed DOI PMC

Bilgin D.D., Liu Y., Schiff M., Dinesh-Kumar S.P. P58IPK, a Plant Ortholog of Double-Stranded RNA-Dependent Protein Kinase PKR Inhibitor, Functions in Viral Pathogenesis. Dev. Cell. 2003;4:651–661. doi: 10.1016/S1534-5807(03)00125-4. PubMed DOI

Hammond R.W., Zhao Y. Modification of tobacco plant development by sense and antisense expression of the tomato viroid-induced AGC VIIIa protein kinase PKV suggests involvement in gibberellin signaling. BMC Plant Biol. 2009;9:108. doi: 10.1186/1471-2229-9-108. PubMed DOI PMC

De Vleesschauwer D., Xu J., Höfte M. Making sense of hormone-mediated defense networking: From rice to Arabidopsis. Front. Plant Sci. 2014;5:611. doi: 10.3389/fpls.2014.00611. PubMed DOI PMC

Xia C., Li S., Hou W., Fan Z., Xiao H., Lu M., Sano T., Zhang Z. Global Transcriptomic Changes Induced by Infection of Cucumber (Cucumis sativus L.) with Mild and Severe Variants of Hop Stunt Viroid. Front. Microbiol. 2017;8:2427. doi: 10.3389/fmicb.2017.02427. PubMed DOI PMC

Bellés J.M., Garro R., Fayos J., Navarro P., Primo J., Conejero V. Gentisic Acid as a Pathogen-Inducible Signal, Additional to Salicylic Acid for Activation of Plant Defenses in Tomato. Mol. Plant-Microbe Interact. 1999;12:227–235. doi: 10.1094/MPMI.1999.12.3.227. DOI

Glazebrook J. Contrasting Mechanisms of Defense Against Biotrophic and Necrotrophic Pathogens. Annu. Rev. Phytopathol. 2005;43:205–227. doi: 10.1146/annurev.phyto.43.040204.135923. PubMed DOI

Yang C.-J., Zhang C., Lu Y.-N., Jin J.-Q., Wang X.-L. The Mechanisms of Brassinosteroids’ Action: From Signal Transduction to Plant Development. Mol. Plant. 2011;4:588–600. doi: 10.1093/mp/ssr020. PubMed DOI

Wang Y., Shibuya M., Taneda A., Kurauchi T., Senda M., Owens R.A., Sano T. Accumulation of Potato spindle tuber viroid-specific small RNAs is accompanied by specific changes in gene expression in two tomato cultivars. Virology. 2011;413:72–83. doi: 10.1016/j.virol.2011.01.021. PubMed DOI

Bhattacharyya D., Gnanasekaran P., Kumar R.K., Kushwaha N.K., Sharma V.K., Yusuf M.A., Chakraborty S. A geminivirus betasatellite damages the structural and functional integrity of chloroplasts leading to symptom formation and inhibition of photosynthesis. J. Exp. Bot. 2015;66:5881–5895. doi: 10.1093/jxb/erv299. PubMed DOI PMC

Attaran E., Major I.T., Cruz J.A., Rosa B.A., Koo A.J.K., Chen J., Kramer D.M., He S.Y., Howe G.A. Temporal Dynamics of Growth and Photosynthesis Suppression in Response to Jasmonate Signaling. Plant Physiol. 2014;165:1302–1314. doi: 10.1104/pp.114.239004. PubMed DOI PMC

Roitsch T., Berger S., Sinha A.K. Plant physiology meets phytopathology: Plant primary metabolism and plant–pathogen interactions. J. Exp. Bot. 2007;58:4019–4026. doi: 10.1093/jxb/erm298. PubMed DOI

Bolouri Moghaddam M.R., Van den Ende W. Sweet immunity in the plant circadian regulatory network. J. Exp. Bot. 2013;64:1439–1449. doi: 10.1093/jxb/ert046. PubMed DOI

Verchot J. The ER quality control and ER associated degradation machineries are vital for viral pathogenesis. Front. Plant Sci. 2014;5:66. doi: 10.3389/fpls.2014.00066. PubMed DOI PMC

Ye C., Dickman M.B., Whitham S.A., Payton M., Verchot J. The unfolded protein response is triggered by a plant viral movement protein. Plant Physiol. 2011;156:741–755. doi: 10.1104/pp.111.174110. PubMed DOI PMC

Fusaro A.F., Correa R.L., Nakasugi K., Jackson C., Kawchuk L., Vaslin M.F.S., Waterhouse P.M. The Enamovirus P0 protein is a silencing suppressor which inhibits local and systemic RNA silencing through AGO1 degradation. Virology. 2012;426:178–187. doi: 10.1016/j.virol.2012.01.026. PubMed DOI

Kim M.Y., Oglesbee M. Virus-heat shock protein interaction and a novel axis for innate antiviral immunity. Cells. 2012;1:646–666. doi: 10.3390/cells1030646. PubMed DOI PMC

Alam S.B., Rochon D. Cucumber Necrosis Virus Recruits Cellular Heat Shock Protein 70 Homologs at Several Stages of Infection. J. Virol. 2016;90:3302–3317. doi: 10.1128/JVI.02833-15. PubMed DOI PMC

Di Serio F., De Stradis A., Delgado S., Flores R., Navarro B. Cytopathic Effects Incited by Viroid RNAs and Putative Underlying Mechanisms. Front. Plant Sci. 2013;3:288. doi: 10.3389/fpls.2012.00288. PubMed DOI PMC

Gorovits R., Czosnek H. The Involvement of Heat Shock Proteins in the Establishment of Tomato Yellow Leaf Curl Virus Infection. Front. Plant Sci. 2017;8:355. doi: 10.3389/fpls.2017.00355. PubMed DOI PMC

Mohr I., Sonenberg N. Host Translation at the Nexus of Infection and Immunity. Cell Host Microbe. 2012;12:470–483. doi: 10.1016/j.chom.2012.09.006. PubMed DOI PMC

Liu B., Qian S.-B. Translational reprogramming in cellular stress response. Wiley Interdiscip. Rev. RNA. 2014;5:301–315. doi: 10.1002/wrna.1212. PubMed DOI PMC

Mitsuda N., Ohme-Takagi M. Functional analysis of transcription factors in Arabidopsis. Plant Cell Physiol. 2009;50:1232–1248. doi: 10.1093/pcp/pcp075. PubMed DOI PMC

Li X., An M., Xia Z., Bai X., Wu Y. Transcriptome analysis of watermelon (Citrullus lanatus) fruits in response to Cucumber green mottle mosaic virus (CGMMV) infection. Sci. Rep. 2017;7:16747. doi: 10.1038/s41598-017-17140-4. PubMed DOI PMC

Sun Y., Fan M., He Y. Transcriptome Analysis of Watermelon Leaves Reveals Candidate Genes Responsive to Cucumber green mottle mosaic virus Infection. Int. J. Mol. Sci. 2019;20:610. doi: 10.3390/ijms20030610. PubMed DOI PMC

Matoušek J., Piernikarczyk R.J.J., Dědič P., Mertelík J., Uhlířová K., Duraisamy G.S., Orctová L., Kloudová K., Ptáček J., Steger G. Characterization of Potato spindle tuber viroid (PSTVd) incidence and new variants from ornamentals. Eur. J. Plant Pathol. 2014;138:93–101. doi: 10.1007/s10658-013-0304-6. DOI

Bernad L., Duran-Vila N. A novel RT-PCR approach for detection and characterization of citrus viroids. Mol. Cell. Probes. 2006;20:105–113. doi: 10.1016/j.mcp.2005.11.001. PubMed DOI

Hataya T., Hikage K., Suda N., Nagata T., Li S., Itoga Y., Tanikoshi T., Shikata E. Detection of Hop Latent Viroid (HLVd) Using Reverse Transcription and Polymerase Chain Reaction(RT-PCR) Jpn. J. Phytopathol. 1992;58:677–684. doi: 10.3186/jjphytopath.58.677. DOI

Li B., Dewey C.N. RSEM: Accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinform. 2011;12:323. doi: 10.1186/1471-2105-12-323. PubMed DOI PMC

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

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

Zhao S., Guo Y., Sheng Q., Shyr Y. Advanced Heat Map and Clustering Analysis Using Heatmap3. BioMed Res. Int. 2014;2014 doi: 10.1155/2014/986048. PubMed DOI PMC

Livak K.J., Schmittgen T.D. Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2−ΔΔCT Method. Methods. 2001;25:402–408. doi: 10.1006/meth.2001.1262. PubMed DOI

Štajner N., Cregeen S., Javornik B. Evaluation of reference genes for RT-qPCR expression studies in hop (Humulus lupulus L.) during infection with vascular pathogen verticillium albo-atrum. PLoS ONE. 2013;8:e68228. doi: 10.1371/journal.pone.0068228. PubMed DOI PMC

Conesa A., Götz S. Blast2GO: A comprehensive suite for functional analysis in plant genomics. Int. J. Plant Genom. 2008;2008:619832. doi: 10.1155/2008/619832. PubMed DOI PMC

Kanehisa M., Goto S., Furumichi M., Tanabe M., Hirakawa M. KEGG for representation and analysis of molecular networks involving diseases and drugs. Nucleic Acids Res. 2010;38:D355–D360. doi: 10.1093/nar/gkp896. PubMed DOI PMC

Supek F., Bošnjak M., Škunca N., Šmuc T. REVIGO Summarizes and Visualizes Long Lists of Gene Ontology Terms. PLoS ONE. 2011;6:e21800. doi: 10.1371/journal.pone.0021800. PubMed DOI PMC

Mao X., Olyarchuk J.G., Wei L., Cai T. Automated genome annotation and pathway identification using the KEGG Orthology (KO) as a controlled vocabulary. Bioinformatics. 2005;21:3787–3793. doi: 10.1093/bioinformatics/bti430. PubMed DOI

Thimm O., Bläsing O., Gibon Y., Nagel A., Meyer S., Krüger P., Selbig J., Müller L.A., Rhee S.Y., Stitt M. mapman: A user-driven tool to display genomics data sets onto diagrams of metabolic pathways and other biological processes. Plant J. 2004;37:914–939. doi: 10.1111/j.1365-313X.2004.02016.x. PubMed DOI

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