Study On Physiological Characteristics And Differential Proteomics Of Pollination And Fertilization In’Tianyuanhong’ Kiwifruit

The all red Actinidia arguta’Tianyuanhong’were studied by field investigation or using fluorescence paraffin technique, to explore the factors that caused its high fruit drop rate. At the same time, Stigmatic and stylar proteins play a pivotal function in plant reproduction during pollination and fertilization. We compared kiwifruit(Actinidia. arguta) Stigmatic and stylar proteins before and10h after pollination. Several specific proteins were identified using fluorescence differential in-gel electrophoresis (DIGE), Matrix-Assisted laser Desorption/Ionization Time of Flight Mass Spectromety (MALDI-TOF/MS) and bioinformatic technology. We also observed the growth of kiwifruit pollen tubes in the style by fluorescence microscope. The main results were as follows:1. The study included flower structure, floral dynamic and life, bloom temperature, stigma receptivity, effective pollination period and embryogenesis of the all red Actinidia arguta’Tianyuanhong’.(1) Female flower branches include the following five types:Stigma of Tianyuanhong’ was dry, it was long cylindrical shape and had a crack ditch. Flowering process is divided into the following five stages:calyx cracking time, calyx large cracking time, petal color change, large flower buds and flowering time.’Tianyuanhong’flowers were neat in steady temperature, its life span of one single flower was approximately1-2days.’Tianyuanhong’receptivity is strong during1st-2nd day and begins to decline during3rd-5th day, and stigma com pletely lose its receptivity on6th day.’Tianyuanhong’ effective pollination period was approximately1-2days. (2) Double fertilization is the type of the Premitotic gametogamy. A sperm and egg cell fusion formation of homozygous; another sperm and secondary Primary endosperm formation of a nuclear fusion.(3) The development of embryo belongs to solanad type. The fertilized egg usually underwent a rest Period about3days before embarking on its first division. And the first division of the zygote took Place after the division of Primary endosperm nucleus. After the club-shaped embryo, the globular embryo, the fish-torpedo and the heart-shaped embryo, the cotyledon-shaped embryo developed when the fruit became ripe. At the Early globular embryo stage, the suspensor was well developed. From the late globular to early heart-shaped embryo stage, the suspensor grew to its minimum length.(4) The endosperm development of’Tianyuanhong’was cellular type. The Primary endosperm nucleus division was often Prior to zygote, and it divided quickly. The endosperm cells surrounding the embryo were disintegrated during the development of embryo.(5) flowering stage, effective pollination period and stigma receptivity with short time are the main factors that affect the female success of’Tianyuanhong’.2. Fluorescence microscope and scanning electron microscopy were used to observe pollen germination and pollen tube growth, Potassiam antimonate was used to localize calcium in stigma and style tissue of’Tianyuanhong’ before and after pollination.(1)1h after pollination, pollen tube grew through stigma surface, and got to basal style10h after pollination.(2) Receptive surface of stigma, Cuticle organelles were rich in calcium granules before and after pollination, but no-receptive surface of stigma, organelles had little calcium granules.(3) Top style had little calcium, but basal style had more calcium before and1h after pollination. The calcium contents were briefly identical in top and basal style10h after pollination.(4) The calcium distributed in plasma membrane of top style before and after pollination, in intercellular gap of basal style before pollination, in plasma membrane of basal style1h after pollination, and in plasma membrane and endoplasmic reticulum of basal style10h after pollination. Stigma and style tissue of’Tianyuanhong’were rich in calcium before and after pollination. Calcium of style showed the gradient distribution before and1h after pollination, it weakened or disappeared10h after pollination. 3. Compared with kiwifruit (Actinidia. arguta) stylus and ovary proteome before and after pollination. Several specific proteins were identified using fluorescence microscope, paraffin technique, differential in-gel electrophoresis (DIGE), Matrix-Assisted laser Desorption/Ionization Time of Flight Mass Spectromety (MALDI-TOF/MS) and bioinformatic technology.(1) According to the DIGE profiles, We gained the two-dimensional gel electrophoresis, which had high resolution ratio and more protein spots proteome.(2) We picked24differential protein spots (ratio>2.5) by DeCyder6.5(Amersham Bioscience), and identified18differential proteins through MALDI-TOF/MS. Of these proteins,8belong to the actinidin family, and2were identified as being the same protein. Among the10significantly different protein spots,5were up-regulated while5were down-regulated, compared with stigmatic and stylar proteins before pollination. Most of the identified proteins play an important role during kiwifruit polliantion. Their putative functions in these processes are discussed.(3) About1500protein points were detected by DeCyder6.5(Amersham Bioscience), and55different proteins were expressed at ovary proteins before and120h after pollination. Thirteen protein spots (ratio>2.0) were identified through MALDI-TOF/MS. Of these proteins,8belong to6kinds of proteins. Among the6significantly different protein spots,5were up-regulated while1were down-regulated. Compared with differences ovary proteins expression before and after Fertilization, we can filter out the double fertilization of ovary proteins.