Functionalanalysis Of PhDof16 In Floralorgans Of Petunia

Petunia(Petunia hybrida Vilm.)Is a herbaceous flower of the Petunia Juss.The main feature is the rich color of flowers and the longer flowering period.The genetic background of petunia is clear,the flower organs are large for easy observation and genetic transformation is easy,so it is often used for flower organ research.The pre-study of the our group discovered an early anther specific expression gene Ph GRP,and its over-expression will eventually lead to stamen abortion.Subsequently,the research group used the bait vector constructed by Ph GRP specific promoter to screen the library and then obtained a PhDof16 transcription factor.In order to identify the function of PhDof16 gene in anther development of petunia,this study carrys out subcellular localization of PhDof16 gene,and construct an interference vector and a over-expression vector.Phenotype observation,cytological analysis and expression analysis of the transgenic plants are also used.1.Through the subcellular localization test we find the PhDof16 gene is expressed inthe nucleus.2.The interference vector 35S::i PhDof16 is constructed and is transformed into Petunia 'W115' by agrobacterium.Finally we get 9 positive plants by PCR detection in 18 resistant plants.Observing the transgenic lines shows that the anther and pistil of line 4 become smaller and shorter,and the crown width of some plants also became smaller;some small and anomalous pollen grains appear in lines 4 and 6;the germination rate decrease overall;the number of pollen grains in a single anther and the number of seed in a single pod of lines 4 and 6 decrease significantly;quantitative PCR results shows the PhDof16 gene expression in lines 4 and 6 reduce obviously.3.The over-expression carrier 35S::PhDof16 is constructed.We obtain 10 positive plants in 15 resistant plants by the same method.Observing the transgenic lines shows that line 5 has a small amount of pollen grains that is significantly larger than the wild type.The germination rate is also significantly lower than the wild type;the number of single anther pollen grains of lines 5 and 16 is lower than the wild type.We find that the surface of the pollen grains is thicker and closer than the wild type by scanning electron microscope.Quantitative PCR detection result shows that the expression level of PhDof16 gene in line 5 is significantly higher than the wild type.The above results preliminarily indicate PhDof16 may have an effect on the development of stamen and corolla.