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Deciphering Regulating Mechanism Of Height And Tiller Development In Tall Fescue Based On Transcriptome Profiling And The Function Of FaGA2ox4 Gene

Posted on:2022-09-23Degree:MasterType:Thesis
Country:ChinaCandidate:R XingFull Text:PDF
GTID:2543307133490044Subject:Grass science
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Tillers and plant height are important agronomic traits for grasses.Identification of genes controlling tillers and plant height is of great significance for molecular assisted breeding of forage and turfgrass.In this study,two varieties of tall fescue(Festuca arundinacea)’Kentucky-31’and’Regenerate’were used as materials to construct transcriptome maps and explore the differentially expressed genes(DEGs)regulating growth and development of crown.In addition,FaGA2ox4 was heterogeneously transferred into rice(Oryza sativa)to explore the effects of FaGA2ox4 on plant height and tiller phenotype,as well as its role in response to drought stress.The experimental results are as follows:1.Transcriptomic sequencing was performed based on Illumina Hi Seq 2500×Miseq300,and the obtained data were assembled by de novo and a total of 77872 unigenes were obtained.A total of 1573 up-regulated DEGs and 1441 down-regulated DEGs were detected in Regenerate compared with Kentucky-31.GO enrichment analysis found that DEGs were mainly enriched in biological processes such as cell,cell component and macromolecular complex assembly.A total of 42 differentially expressed transcription factors were noted in DEGs,and mainly belonged to TCP,WRKY and ARF families.The DEGs were associated with 8 plant hormones,including Indole-3-acetic acid(IAA),Cytokinin(CTK),Abscisic acid(ABA),Gibberellin(GA),Ethylene(ETH),Brassinosteroids(BR),Salicylic acid(SA)and Jasmonic acid(JA).The expression patterns of DEGs verified by real-time PCR were consistent with RNA-Seq,indicating the RNA sequencing results were accurate.2.The tissue expression analysis of FaGA2oxs genes(FaGA2ox1,FaGA2ox2,FaGA2ox3,FaGA2ox4,FaGA2ox7,FaGA2ox8,FaGA2ox9 and FaGA2ox10)showed that these genes were expressed in all tissue parts.However,the expression level was the highest in leaves.Tall fescue plants were treated under low temperature,high temperature,salt and drought conditions respectively,and the expression level of theses degrading enzyme genes changed under different stress conditions.The results showed that FaGA2oxs genes were involved in plant stress response mechanisms.3.The phenotypes of over-expressed FaGA2ox4(Line6,Line8)and WT(Wild type)rice plants at different time points were analyzed statistically,and the contents of endogenous hormones were determined by high performance liquid chromatography-tandem mass spectrometry.Over-expression of FaGA2ox4 gene reduced the contents of GA53,GA8and GA20in rice plants,which resulted in GA-deficient phenotypes in rice plants,such as dwarfing,increased tillers,short and wide leaves.The 10μM GA3solution was sprayed once every 7 days for each line.The experimental results showed that exogenous GA3could compensate the GA-deficient phenotype of the over-expressed FaGA2ox4 rice plants.4.By using 15%polyethylene glycol(PEG)to simulate drought treatment on21-day-old rice plants,experiment results showed that the leaf chlorophyll content,relative water content and photosynthetic rate of transgenic lines were higher than those of WT,and the relative electrolyte leakage of transgenic lines were lower than those of WT.The content of ABA in Line6 was significantly higher than that in WT.The expression level analysis of chlorophyll synthesis and metabolism(Os CAO1,Os SGRL),GA(OsGA3ox1,OsKS1)and ABA(OsNCED3)synthesis and drought stress responsive genes(Os NHX1)under control and PEG treatment were detected by q RT-PCR.Experimental results showed that the expression levels of these genes were different between transgenic lines and WT,indicating that these genes may be involved in the molecular mechanism of FaGA2ox4mediated drought response..In conclusion,this study enriched the transcriptome sequence resources of tall fescue,compared the molecular mechanism differences between’Kentucky-31’and’Regenerate’in growth and development,especially in tiller and plant height development,and preliminarily obtained candidate genes controlling plant height and tiller development,which provided theoretical support for further gene function and molecular breeding research.It was confirmed that FaGA2ox4 could affect plant height and tillering,and also participate in plant drought stress responses.
Keywords/Search Tags:tall fescue, RNA-seq, gibberellin 2-oxidases 4, tillers, height, drought tolerance
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