Preliminary Function Analysis Of The Flower Development Related Genes And Transformation In Platanus Acerifolia Willd

Because of its broad crown, fast growth, wide adaptability, good tolerance to pruning, and high resistance to environmental pollution, Platanus acerifolia willd is called "the king of street tree", and it becomes as one of the most important landscaping tree species. However, its pollen and seed hairs disperse widely, which not only pollute the environment, but also seriously affect human health. At present, there is very few research on the pollution of the pollen and seed hairs onabroad. Many researchers have carried out a lot of studies on this subject in China, but have not resolved the problem completely.According to the present situation, as well as the latest progress of the key flowering genes and MADS-box genes, the expression of floral development related endogenous genes in P. acerifolia by genetic engineering would be disturbed, so that flowering and floral organ development of P. acerifolia can be interfered, which would lay the foundation for cultivating less or even non-seed hair about it. A t the same time, in order to analyze the preliminary function of the floral development related genes in P. acerifolia, the sence and anti-sence genes were introduced respectively into tobacco by Agrobacterium tumefaciens-mediated transformation, so that the phenotype variations of transgenic plants can be anlalyzed, including flowering time, size, floral organ development, fruit development and so on. Furthermore, in order to improve the efficiency of Agrobacterium-mediated transformation of P. acerifolia, some factors about it were optimized, and some target genes of P. acerifolia were transformed. The main results as following:1. Anti-sense plant expression vectors of PaLFY, PaFUL, PaAP3, PaSEP1, PaSEP3 and PaLFY-FUL were constructed.2. Sense and Anti-sense plant expression vectors of PaLFY、PaFUL、PaAP3、PaSEP1、PaSEP3 and anti-sense PaLFY-FUL were introduced respectively into tobacco by Agrobacterium tumefaciens-mediated transformation. The analysis of PCR showed that these heterologous genes were successfully introduced into tobacco. Comparing with the control:(1) Sense PaLFY transgenic tobacco showed the phenotype of early flowering, reduced number of leaves and dwarf plant size, but antisense PaLFY transgenic tobacco didn’t show obvious phenotype. It indicated that PaLFY maybe prompt early flowering.(2) Sense PaFUL transgenic tobacco showed the phenotype of early flowering, dwarf plant size, and significantly reduced number of leaves. The strong expression lines can generate many lateral inflorescences, which would generate secondary inflorescences, most inflorescence of lateral branches are higher than that of the stem; Antisense PaFUL transgenic tobacco showed delayed flowering time and increased number of leaves, but plant height internode, the number of leaves, leaf length and width had not obvious phenotype changes comparing with the controls.(3) Sense PaAP3 and antisense PaAP3 transgenic tobacco didn’t show obvious phenotype changes either vegetative growth or flowering time. However, some lines of sense PaAP3 transgenic tobacco showed wrinkled corolla rim, another had lacerated corolla, and the others even had auxiliary petal; Some antisense PaAP3 transgenic lines showed the phenotype of shorter corolla with exposed stamen and stigma, or shorter corolla and stamen, with exposed stigma. It indicated that PaAP3 maybe participate in the development regulation of petal and stamen.(4) Sense PaSEP1、PaSEP3 transgenic tobacco showed the phenotype of early flowering, dwarf plant size, shorter internodes, smaller leaf shape and significantly reduced number of leaves. The strong expression lines can even generate many lateral inflorescences, which would generate secondary inflorescences, and most inflorescence of lateral branches is higher than that of the stem. Some sense PaSEP3 transgenic tobacco especially generate single terminal flower. However, antisense PaSEP1、PaSEP3 transgenic didn’t show obvious phenotype changes in plant height, internode, the number of leaves during the flowering conversion, leaf length and width,as well as the first flowering time.(5) Double antisense PaLFY-FUL transgenic tobacco didn’t show obvious phenotype changes in plant height, internode, and the number of leaves during the flowering conversion, leaf length and width, as well as the first flowering time comparing with the controls. To further analyze the genetic stability of these transgenic tobaccos, the seeds of To generation were seeding and T1 generation plants are at vegetation growth up to present.3. To improve the efficiency of Agrobacterium tumefaciens-mediated trans-formation of P. acerifolia, some lines with the ability of regeneration and differentiation were screened from 236 lines firstly.2 superior lines, C19 and Q55 were primarily slected as the subsequent experiment material, whose regeneration frequency can reach up to 70%. On the other hand, the plant culture medium, proliferation of in vitro shoots and differentiation of resistant buds, were adjudted. MS+BA0.4mg/L+NAA0.05 mg/L and MS+BA5.0 mg/L+ IBA0.5 mg/L is respectively used as proliferation medium and differentiation medium.4. Some resistant lines of P. acerifolia have been obtained by Agrobacterium tumefaciens-mediated transformation, including sense PaFT, PaSEP3、antisensePaLFY、antisense PaFUL、antisensePaFUL-LFY. But the analysis of PCR showed that they were false positive. The possible reason is either the time of co-cultivation has been reduced, or the time of delay-cultivation has been increased. The resistant lines of BpFUL-Barnase are at the stage of strong seedling up to the present, and they haven’t been analyzed.