The Study On Molecular Mechanism Of Cucumber Parthenocarpy Based On Transcriptom

Cucumber (Cucumis sativus L.) is an important vegetable crops widely cultivated around the world, plays an important role in the production and consumption. Due to the low-temperature, low light and high humidity and other factors in the protected cultivation result in poor pollination and fertilization, resulting in abnormal fruit development, thus affecting the economic efficiency of agriculture. Parthenocarpy is an important trait and one of the main breeding objectives in protected cultivation for cucumber. China has rich parthenocarpic cucumber germplasm resources, but many facilities cultivars of cucumber are lack of the parthenocarpic trait, the fruit development affected by the environment, which greatly affected the production and quality of cucumber. Thus, the study on cucumber parthenocarpy, and exploring genes related with parthenocarpy has an important significance for improving cucumber cultivars. In this paper, the genes related with cucumber parthenocarpy were found by the RNA-Seq technology with the cucumber parthenocarpic variety’EC1’and non-parthenocarpic variety’8419s-1’as test materials. Then gene transient expression method was used for rapid functional verification about the part candidate genes of cucumber parthenocarpy. Based on the differentially expressed genes involved in cucumber parthenocarpic fruit development from the analyses of RNA-Seq, two auxin receptor genes CsTIR1and CsAFB2were chosen for the analyses of expressional level in different cucumber organs, and their response to exogenous hormones were studied. In addition, the CsTIR1were over-expressed in tomato for functional verification. The temporal and spatial expression expressional patterns of CYCD genes were also studied for understanding of cell division in the early stage of cucumber parthenocarpic fruit. The main results are as following:1. Using RNA-Seq technology to explore the molecular mechanism of cucumber parthenocarpic fruit set and the preliminary function of genes related with parthenocarpyIn present study, RNA-Seq technique was used to collect the transcriptome data of cucumber fruits2days after anthesis for screening of parthenocarpy genes. Differentially expressed genes (DEGs) were analyzed in both parthenocarpic and pollinated cucumber fruits contrasting with abortive fruits. Trough comparative analyses, a total of1339genes associated with parthenocarpic fruit development were obtained, including759genes up-regulated and580down-regulated. Expression patterns of phytohormone related DEGS supported that phytohormones are key factors for fruit setting. Also selected12of them involved in fruit development process were analyses in early developmental parthenocarpic fruit, and the results indicated that the selected genes play an important regulatory role in parthenocarpic fruit development.The transient expression of gene in plant tissues is a very good method for preliminary research of gene function. In the present study, we used the method of Agrobacterium-mediated transient expression to analysis cucumber parthenocarpic fruit-specific expression of gene function. The results showed that the exogenous gene introduction after10days, Cucumber ovary appeared the following states:expansion fruit, the stopped development ovaries but not wilt and wilt ovaries. GUS assay results in wilting ovary did not carry exogenous gene of Agrobacterium infection to cucumber ovary tissue, so the exogenous genes were not been transformed in ovary tissue. Studies have shown that the selected10genes to stimulate parthenocarpic fruit and this technology can also be used for the function analysis of cucumber fruit characters genes.2. Expression level and function analyses of auxin receptor genes in cucumber parthenocarpic fruit developmentIn our study, CsTIRl and CsAFB2were isolated from RNA-Seq analysis, it were highly expressed in ovaries and leaves and their transcript levels decreased in parthenocarpic and pollinated fruits, but continuously up-regulated in non-parthenocarpic and unpollinated fruits, indicating that down-regulation of CsTIRl and CsAFB2may be necessary for cucumber fruit set and development. The transcript levels of CsTIRl and CsAFB2were significantly induced in leaves by NAA,6-BA, GA3, ABA, and ethephon. The expression levels were up-regulated by ABA and ethephon. These results suggest that CsTIRl and CsAFB2may be involved in abiotic stress response. Thus, CsTIRl and CsAFB2may be important regulators during cucumber fruit development.Auxin receptor TIR1plays an important role in regulating the growth and development of plants. In order to study the function of the cucumber CsTIR1, it was transformed into tomato by Agrobacterium-mediated transformation. The results showed that the transgenic tomato lines overexpressing CsTIR1exhibit parthenocaarpy and dwarf. This proves that the CsTIR1gene may be a key regulator in parthenocarpic fruit development.3. Genome-wide identification of D-type cyclins gene family from cucumber and analysis of their expression patterns in fruit developmentCell division plays an essential role in early fruit development. Plant D-type cyclins (CYCDs) are important regulators in the cell division. However, the knowledge regarding their involvement in early developmental stage of cucumber fruit remains limited. To address this, the cucumber CYCD family genes and their structural characteristics were identified, and then their expressional patterns in early fruit development were analyzed from the anthesis day (0d) to8days after anthesis (8d). The result showed that the cucumber genome contains13different CYCD genes which have been named according to identity percentages of the corresponding orthologs in Arabidopsis and poplar (Populus trichocarpd). The genomic organization of each subgroup cucumber CYCD showed the structure similar to their orthologs in Arabidopsis and poplar. The expression levels of cucumber CYCD genes were investigated in early stage of fruit development under different treatments, including:natural parthenocarpic, pollination, induced parthenocarpic with CPPU treatment. The highest expression level of most of CYCDs appeared at4DAA in natural parthenocarpic and pollinated fruit. Moreover, the expression patterns of most CYCD genes in natural parthenocarpic fruit were nearly coincident with in pollinated fruit, but different from induced parthenocarpic fruit. Collectively, these results will provide molecular insights into the CYCDs which are contributing to the formation of cucumber parthenocarpic fruit.