Transcriptome And Small RNA Analysis Of Seed Germination And Gibberellin Response In Rice

Generally,seed germination is a rapid transition process from dormancy to seedling growth,which starts a new round of life cycle for plants.Rice is one of the most important crops in the world,it is also a model plant for Single leaf plant.As genome sequencing technology and omics" developed,transcriptome sequencing become a new tool for seed germination study.Seed imbibition injury is an important factor affecting Direct seeding rice seeding.To determine the regulators that are involved in stress responses during seed germination,dynamic transcriptome analyses were conducted at dry(0 h),imbibed(24 h)and germinated(48 h)stages of elite japonica landrace Jiucaiqing using RNA sequencing method in this study.Results showed that the differentially expressed genes(DEGs)were pronounced in the early stage(0-24 h)than the late stage(24-48 h);1,452 transcripts were differentially expressed include 648 up-regulated and 804 down-regulated in the early stage.got 625 transcripts include 230 up-regulated and 395 down-regulated in the late stage.Gene ontology and MapMan analyses confirmed that 391 and 164 DEGs at the early and late stage respectively involved in stress responses pathway.These DEGs included the abiotic stress-,hormone-,peroxidases-,signaling-,transcription-,proteolysis-and cell wall-related genes.Detailed dissection indicated that the mainly heat stress-related DEGs and many auxins-,abscisic acid-and ethylene-related DEGs were involved in stress responses during seed germination.Nearly all the peroxidases-related DEGs and the majority of signaling-related DEGs,e.g.calcium-binding proteins,were up-regulated with seed germination.Several transcription factors of bZIP and MYB,ethylene responsive element binding protein family(ERF),and heat stress transcription factor family(HSF),and the proteolysis-related DEGs,e.g.ubiquitin-related proteins,related to stress responses were significantly regulated at the early stage.Many genes involved in cell wall synthesis,degradation and modification were up-regulated during seed germination.These stress responsive genes identified in this study may play an important role in rice seed resistance to imbibition.Small RNAs are 20-24 nucleotides non-coding RNAs.Small RNA-mediated post-transcriptional regulation plays an important role in plant growth and development as well as responses to different environmental stresses(1-5).The number of small molecule RNA in plants is very large,and has a wide range of species,high-throughput sequencing technology has greatly accelerated their discovery process.MicroRNA belongs to endogenous gene regulatory system,It is widely distributed in plant genome and plays an important role in plant growth,development,signal transduction and response to environmental stimuli.In recent years,sequencing of small RNA libraries from different rice cultivars or tissues has severally been reported where a large number of rice miRNAs were identified.Despite this,there are few reports on the study of response of rice small RNAs to phytohormones.GAS play significant regulatory roles in rice.Till now,it is still unclear of how GAs participates in the regulation of the growth and development of rice via small RNA.To study the effect of Gibberellin on small and RNA during rice growth and development,We sprayed gibberellin GA3 and GA3 inhibitor PAC in rice 3 leaf stage,respectively,and the total RNA was extracted for small RNA high-throughput sequencing.The results showed that the Expression level of 26,36 miRNA and 607,939 sRNA(except MicroRNA and its isoforms)changed after after GA3 and PAC treatments.After target prediction and degradom verification,we obtained a total of 63 eligible small RNAs and their corresponding 153 target genes.The GA responsive small RNA regulatory network diagram contains 19 miRNA and 102 sRNAs that are screened by differential expression and are verified by the degradation group and 43 GO term(p<0.01).The selected subnetworks,related to the reproductive development,transcription,metabolic,catalytic activity and transferase activity,were further constructed and discussed.The result shows:Developmental subnetwork related miRNA contains reproductive related miRNA,In addition,the developmental pathway is also related to miRNA1861N,miRNAN58 and miRNA528-5P;MiRNAnovel9 is involved in transcription factor activity,transcriptional regulatory activity,and transport activity GO term.the major target genes of miRNA;56L-5P and miRNA169A are involved in transcription factor activity,MiRNA160B-5P,miRNA160C-5P,miRNA160D-5P,readll,read12 and read13 also participate in transcription regulation and transcription factor activity GO term through their specific target genes.MiRNA528-5P has 5 target genes involved in the transport of the active GO term;Most of the target genes of miRNA156L-5P,miRNAnovel9,miRNA169A,miRNA393A,miRNA393B-5P,miRNA160B-5P,miRNA160C-5P,miRNA160D-5P and miRNA2055 are involved in the metabolic pathway of rice;miRNA2055 related to transferase activity GO term,miRNA528-5P?miRNAnovel58?miRNA393A?miRNA393B-5P?read6 related to catalytic activity GO term.The small RNA regulatory network reveals that most of the small RNAs are involoved in growth,development and stress responses in GA signaling.