Comparative Transcriptome Analysis Of Stamen Petalody In Tree Peony

Tree peony,a perennial shrub plant,which belongs to the genus Paeonia L.Section Moutan DC.(Paeoniaceae),is one of the most important ornamental flower in worldwide.There are more than 2000 various tree peony cultivars in the world,among which 1200 varieties(60%)exist the phenotype of stamen petalody,so stamen petalody is one of the main reasons for the formation of plenty and peculiar flower patterms in tree peony.Therefore,this study will be valuable for understanding the mechanism of stamen petalody,and has a guiding significance for the breeding and production practice of peony and other ornamental plants.In this study,the traditional varieties 'Er Qiao','Wu Long' and'Yu Louchun' were used for the research,and 12 samples,including the petals,petaloid stamens,stamens in squaring bud stages and petals in eoloring bud stages were used to eonducted a transcriptome analysis.A preliminary analysis of the mechanism of stamen petalody and flower organ formantion were conducted,and the phylogenetic analysis of MAD S-box gene family,which has a decisive effect on the formation of peony floral organ were also conducted.The results were as follows:1.Transcriptome sequencing.Three traditional varieties'Er Qiao','Wu Long' and 'Yu Louchun'were selected,12 samples including the petals,petaloid stamens,stamens,in squaring bud stages,and petals in coloring bud stages were used as experiment materials.A total of 118.68 clean reads were obtained from 136.61Gb raw reads by Illumina Hiseq deep sequencing.And got 177312 Unigenes after assembled all samples toget:her.The average length and N50 was 1038 bp and 1664bp,respectively.A total of 102577(57.85%)Unigenes were annotated in NR,NT,Swissprot,COG,KEGG,GO and Interpro 7 functional database,finally.According to the functional annotation results,95152 CDS were detected.2.Comparative transcriptome analysis.EBSeq method was used to analyze the DEGs in squaring bud stages.A total of 2423 DEGs were obtained in petals VS stamens,among which 1181 DEGs were up-regulated and 1242 DEGs were down-regulated.601 DEGs were obtained in petals VS petaloid stamens,among which 436 DEGs were up-regulated and 165 DEGs were down-regulated.And 451 DEGs in petaloid stamens VS stamens,among them 364 DEGs were up-regulated and 87 DEGs were down-regulated.Six down-regulated MADS-box DEGs related to stamens turning into petals were obtained by Gene Ontology and KEGG enrichment analysis:one AGL6 gene and three API genes which control the formation of the petals,one AP3 gene control the formation of the petals and stamens,a SEP genes control the formation of all flower organs.3.Gene expression analysis.Analysis the genes belong to ABC(D)E model based on NR database using the FPKM value showed that,the expression of AP1 and AP2 in the petals was higher than that in the petaloid stamens,and the expression level of API and AP2 was low in the stamens.AP1 and AP2 participate in the role of petals and petaloid stamens identity.The class B genes(AP3 and PI)were expressed highly in the flower organs,regulating the formation of petals,petaloid stamens and stamens.C-class genes(AG and SHP)expressed highly in the stamens,the expression level in petaloid stamens were halved,but hardly expressed in the petals,it controls the development of stamens and petaloid stamens.E-class gene(SEP)was expressed in every organs,regulating the formation of stamens,petaloid stamens and petals.Our results show that the reduction expression of C-class function genes(AG and SHP)and the increased expressed of B-class function genes(AP3 and PI),resulting in stamens converted into petals identity.At the same time,stamen petalody requires the presence of the homeotic A-class genes(AP1 and AP2)and E-class genes(SEP).4.Phylogenetic analysis of tree peony MADS-box gene family.75 MADS-box Unigenes were identified by HMMER research in tree peony transcriptome CDS database.A Maximum Likehood tree containing MADS-box genes from tree peony and Arabidopsis were constructed.According to the homologous function annotations and phylogenetic analysis indicated that,24 of them were type ? genes,while 51 were type ?.Type ? was divided into four subfamilies Ma,Mp,M? and M?.Type ? can be divided into 12 subfamilies in tree peony.A total of 44 genes were ABC(D)E model,Among them,there are 38 genes belongs to MADS-box gene family and 6 genes were AP2.In summary,A high quality and comprehensive transcriptome database was constructed and the key candidate genes involved in stamen petalody were screened in this study.Based on the phylogenetic analysis of the MADS-box gene family,the ABC(D)E model genes of floral organ development were analyzed in tree peony,we inferred that the reduced expression of AG(AGAMOUS)genes gave rise to increased expressed of PI genes results in the phenotype of the stamen petalody.In addition,the regulation model of stamen petalody in tree peony was proposed.This study provides the theoretical basis for the flower formation mechanism of ornamental plants.