Morphology Observation And Study On Regulation Mechanism Of Phytohormones During Floral Sex Differentiation In Persimmon

Persimmon(Diospyros kaki)is a tree belonging to family Ebenaceae and genus Diospyros,which is native to China and it‘s cultivation area and yield are the highest in the world.The flowering habit of persimmon is very complex.Most varieties of persimmon only bear pistillate flowers(pistillate type),while a small number of varieties bear both pistillate and staminate flowers(monoecious type).Additionally,a few varieties bear hermaphroditic flowers in addition to pistillate and staminate flowers(polygamomonoecious type),and a very few varieties just bear male flowers.The lack of male resources strongly restricts hybridization of persimmon.Therefore,it is very important for hybridization of persimmon to induce male resources artificially.To induce male resources,we need firstly to understand the regulatory mechanism of flower sex differentiation in persimmon.Hence,in the present study,male and female floral buds in different developmental stages of monoecious persimmon cultivar ?Zenjimaru‘ were respectively collected and observed through paraffin section method and scanning electron microscope to determine the critical morphological period of floral sex differentiation in persimmon.Subsequently,based on the critical morphological period of floral sex differentiation,male and female flowers collected from three ?Zenjimaru‘ trees with consistent and good growth conditions were used for the analysis of endogenous phytohormone contents through high-performance liquid chromatography-tandem mass spectrometry(HPLC-ESI-MS/MS)to find out the crucial phytohormone regulating sex differentiation.These results will provide theoretical foundation for further study on regulatory mechanism of flower sex differentiation and exploitation of artificial regulation technique.The major results are described as follows:(1)The developmental course of female flowers was basically synchronous with male flowers in ?Zenjimaru‘.The whole development process of both male and female floral buds could be divided into 11 stages ranging from June to May of the next year.There were two morphological key periods about floral sex differentiation in ?Zenjimaru‘.One was mid-June(stage 2)with the initiation of sepal primordia,when male flower formed a cymose structure,while only the terminal flower primordium developed in female flower.The other important developmental phase was mid-April(stage 8)with the initiation of megaspore and microspore,during which stamen primordia in female flowers arrested after the filaments differentiated and the pistil primordia in male flowers also arrested after the stigma and style developed,which led to the formation of unisexual flowers.(2)The embryological characteristics of flower development in ?Zenjimaru‘ was similar to most plants.The anthers were tetradymous and the anther wall consisted of epidermis,endotheca,middle layer and glandular tapetum;meiosis of microspore mother cell was followed by simultaneous cytokinesis and resulted in tetrahedral tetrads;the mature pollen grains were 2-cell type with three germ pores;the giant pollen(unreduced pollen)formed during meiosis of microspore mother cells.The ovary had eight locules with axile placenta and anatropous ovules;the ovule was bitegmic and tenuinucellate with single spore cell;megaspore mother cell formed a linear tetrade through meiosis,and only the chalazal magaspore was the functional one;the development of embryo sac was of polygonum type.(3)On the second critical morphological period of floral sex differentiation(stage 8)in persimmon,the concentrations of IAA,ABA,ZT and JA in female flowers were significantly higher than that in male flowers,while GA3 contents in female flowers were significantly lower than that in male flowers.These data indicated that IAA,ABA,JA and ZT could contribute to the formation of female flowers by accelerating stamen abortion.In particular,the role of IAA and ZT was more obvious,which were the critical regulation phytohormones for the development of female flowers.In the same way,GA3 could promote the formation of male flowers by suppressing the development of pistil primordia,which was the critical regulation phytohormone for the development of male flowers.In addition,the modification effect of these phyhormones happens around stage 8,which is basic accordance with the key differentiation time-point in morphology.(4)The experiments about the spraying of exogenous hormines showed that,ethrel could promote the development of male flowers in persimmon.