Cloning And Functional Analysis Of PsGA20ox,PsBKI1 And PsFabG Genes In Tree Peony

Plant type is one of the important ornamental characters of garden flowers.Endogenous hormones are important aspects of regulating plant architecture,and their synthesis,decomposition and signal transduction regulation affect the establishment of plant architecture.Tree peony is a world famous flower with a wide variety and plays an important role in national life.However,most of the existing tree peony varieties have a single plant type and lack of miniature potted or cut flower varieties.In order to clarify the mechanism of the formation of tree peony plant type traits,explore the key genes regulating plant type traits,and provide technical support for minipot breeding of tree peony,this study screened candidate genes regulating plant type traits by RNA-seq in the early stage of the our research group.Based on this,the cultivar ’Fengdan’ of Paeonia ostii was treated with exogenous hormones.According to the plant phenotype,stem anatomy,endogenous hormone content changes and candidate gene expression,three genes involved in gibberellin synthesis key oxidase,BRI1 kinase inhibitor involved in receiving brassinolide signal reception and mitochondrial fatty acid synthesis key reductase were selected.It is speculated that they may be involved in the regulation of peony plant type traits.The full-length sequences of PsGA20 ox,PsBKI1 and PsFabG were cloned by RACE technology.The physicochemical properties,conservation and evolutionary relationship of homologous sequences of the target gene were analyzed by bioinformatics.The plant overexpression vectors were constructed and the key genes were functionally verified in Arabidopsis thaliana.The main conclusions of this study are as follows :1.The PsGA20 ox,PsBKI1,and PsFabG genes all responded to exogenous hormone treatment-expression levels were up-regulated or down-regulated to varying degrees within a few hours after treatment;the complete CDS of the cloned genes were 1149 bp,873 bp and 972 bp,respectively.The plant overexpression vectors of p Bi121-PsGA20 ox,p Bi121-PsBKI1 and p Bi121-PsFabG were successfully constructed.2.Bioinformatics analysis showed that the PsGA20 ox gene encodes hydrophilic,extracellular,non-secretory proteins,and its encoded protein sequence has a highly conserved region,which is closely related to Camellia sinensis in evolution.The PsBKI1 gene encodes a hydrophilic protein and has the possibility of transmembrane,but it is a non-secretory protein.The overall coefficient of variation of the protein sequence is large and it is not conservative.It is closely related to the Camellia sinensis in evolution.The PsFabG gene encodes a hydrophobic,extracellular protein,which is a non-secretory protein with a conserved region.It is first clustered with Vitis vinifera in evolution,followed by Camellia sinensis.3.It was found that PsGA20 ox could promote the development of lateral roots and fibrous roots of Arabidopsis thaliana and promote it early bolting.At the same time,the color of rosette leaves of Arabidopsis was lighter than that of the control,and the growth of the plant declined in the later stage.The anatomical analysis of the elongation zone at the top of the stem segment showed that the transgenic plants had more dense conducting tissues than the wild type.4.The survival rate of PsBKI1 transgenic Arabidopsis seeds after germination was higher than that of wild-type lines,and the lateral root hairs were dense;At the seedling stage,the number of rosette leaves was more than that of the control,the leaf color was lighter,and the leaf margin had obvious serration.In addition,the plant bolting early,lateral branch development exuberant and the anatomical results showed that the pith cells of PsBKI1 transgenic Arabidopsis stems were larger than those of wild type.5.The seed germination rate of PsFabG transgenic Arabidopsis was low;the plant bolting time was consistent with the control,and the mature plant height was lower than the control.There was no significant change in the number and morphology of rosette leaves,only the leaf color was reddish.Stem anatomy showed that the size of xylem cells in the stem of transgenic plants was significantly larger than that of the control.The results of this study will provide an experimental direction for the functional study of three target genes,and provide an important theoretical basis for the molecular regulation mechanism of peony plant type traits,so as to lay an important experimental foundation for the directional cultivation of peony varieties with different plant types.