Comparative Analysis Of PhAP1 And PhFBP29 Gene Function In Petunia Hybrida

Petunia PhAP1 and PhFBP29 belong to SQUA subfamily of MADS-box gene family.AP1 is a key gene regulating flowering in Arabidopsis thaliana,and it can promote flowering.The sequence and function of AP1 gene in Petunia hybrida have not been reported.Preliminary studies showed that Petunia PhFBP29 could also promote flowering.Petunia is one of the most popular flower bed plants and is a model material of comparative genomics.Gene editing using CRISPR/Cas9 in petunia can provide the basis for other ornamental plants.At the same time,it can improve the characters and ornanmatal value of petunia.In this study,we compared and analyzed the two gene functions by comparing the phenotypic changes of transgenic plants between knockout PhAP1 and PhFBP29 gene mutants and over-expressing PhAP1 and PhFBP29.The PhAP1 gene was cloned from Petunia,and the bioinformatics and transcriptional level of the gene were analyzed.Meanwhile,we separatly constructed vectors,and obtained over-expressing transgenic plant and mutant plant.The phenotype of transgenic plant was observed and the functions of PhAP1 and PhFBP29 genes of Petunia were analyzed.The main results are as follows:1?sequence comparison of PhAP1 and PhFBP29 genes in PetuniaUsing Biosoftware analysis,We found that the maximum ORF frame of the PhAP1 gene was 723bp,and the AP1 protein was composed of 241 amino acids,which was an unstable hydrophilic protein without a transmembrane domain.This protein contains highly conserved domain MADS and a relatively conservative domain K,PhAP1 gene is the MADS-box gene of plant specific MIKC.The AP1 protein consists of 66.39%alpha helices,25.31%random curls and 8.3%extended chain three secondary structures.The secondary structures and physicochemical properties of the PhAP1 and PhFBP29 genes are similar,but the two genes belong to the different branches of the AP1/FUL subfamily.The PhAP1 gene belongs to the euAP1 branch,and the PhFBP29 belongs to the FUL-Like branch.2?Differences in PhAP1 and PhFBP29 gene expression characteristics of Petunia hybridaThe tissue specificity of the PhAP1 gene of Petunia was analyzed.It was said that the PhAP1 gene of Petunia was highly expressed in the stem and flower.The expression level of PhAP1 gene in stem tip varied during developmental stages,and it was the highest in flowering stage,but hardly expression at seedling stage.Although the PhAP1 and PhFBP29 genes were hardly expressed in roots,both were expressed in the flower organs and the stems of Petunia.PhFBP29 gene could expressed in the leaves,but the PhAP1 gene was not expressed in the leaves.3?Preliminary analysis of the function of FBP29 and PhAP1 genes in PetuniaThe T₁ and T₂ generation transgenic Petunia phenotype of overexpressing PhFBP29?521?and overexpressing PhFBP29-SRDX?522?were observed.At the same time,the PhAP1 gene overexpression,knockout PhAP1 and PhFBP29 vectors were constructed,respectivly.PhAP1?PhFBP29 gene knockout mutants and PhAP1overexpressed transgenic plant were obtained.Through Agrobacterium mediated transformation,8 lines mutant M₀ generation Petunia were obtained.The seeds of transgenic Petunia were collected,homozygous mutants of single gene homozygous was obtained from M₁,and double gene homozygous mutants was obtained from hybridization.The flowering time of T₁ generation overexpressing PhFBP29 petunia was earlier than that control Petunia,but the T₁ and T₂ generation overexpressing PhFBP29-SRDX petunia were dwarf,flowering delayed,leaf and leaf epidermis smaller,lateral branches increased,and stem internodes shorter.The flowering time of the overexpressed PhAP1 transgenic T₀ generation Petunia was earlier than control Petunia.The flowering time of knockout PhAP1 M₁ generation mutant plants was shorter than control.Both PhFBP29 and PhAP1 genes can regulate flowering time.PhFBP29 gene plays an important role in the cell size of leaf epidermis and leaf epidermis.It is an important gene controlling leaf development and lateral development.The function of PhAP1 gene in leaf development and floral organ development remains to be studied.