| Lily is a kind of perennial bulb flower with large and colorful flowers. It has been one of the most important fresh cut flowers and is very popular all over the world. But its pattern schema is monotonous.Up to now, most of the wild-type lilies are simple flower. Besides, the problem of pollen contamination in lily will not only affect aesthetic compromises but also shorten its vase life.For a long time, people pay much attention to the shape breeding and long for new cultivars with higher ornamental value, especially double flowers with its noble and plump beauties.However, double flower or no pollen lily is hard to find through natural mutation. If any, they can hardly be used as breeding parents. The development of molecular biology and biotechnology make it possible to solve these problems in a short time. Flower development ABC model is a hot spot nowadays.Studies with model plants of Arabidopsis thaliana and Antirrhinum majus led to a deeper understanding of ABC model of flower development.In this study, we isolated a gene form lily cultivar'seberia' named LLAG homologous to flower development C type genes.We constructed the over expression vector, anti-sense vector and the interference vector. On the one hand, these plasmids were transformed into tobacco for functional verification via agrobacterium-mediated genetic transformation method. On the other hand, they were transformed into petunia expecting some changes in its pattern schema to enrich the variety of petunia and provide supports for obtaining double or pollenless lily at last. The experimental results were as follows:1.After designing primers according to the reported flower development C type gene in lily, we cloned a C type gene named LLAG homologous to LLAG1.It shares a high sequence similarity to the reported flower development C type gene in Lilium longiflorum. Initially, we considered LLAG as a flower development C type gene in Lilium cultivar'seberia'.2.Add appropriate enzyme sites to the cloned LLAG gene, and then, constructed the oversexpression vector and anti-sense vector using pMV vector; constructed the interference vector using pHANNIBAL and pART27 vector. At last, we transformed these vectors into agrobacterium.3.We transformed these vectors into nicotiana tabacum and petunia hybrida. In total,49 pMV-oxLLAG transgenic tobaccos and 12 transgenic petunias were positive; 31 pMV-anti-LLAG transgenic tobaccos and 4 transgenic petunias were positive; 21 pA-LAGRi transgenic tobaccos and 6 transgenic petunias were positive.4.Three different types of homeotic changes occurred in the floral organ of over expression vector transgenic tobaccos. The first type experienced enlarged sepal, the corolla tube opened at the young stage when it was not yet out of the calyx, the shrinkage petals couldn't open completely. Sepal of the second type enlarged more seriously than the first one, apex of the corolla tube was bulging, petals rolled out and couldn't open fully. Changes in type three were the most seriously, apex of the seapl rolled out and the base of the sepal enlarged. Petals reflexed inward and an appendage similar to petal occured at the base of the corolla tube. Scanning electron microscope showed that the cell morphology of the transgenic tobacco was similar to the cell morpholoty of wild-type anthers.RT-PCR demonstrated that exogenous C type genes can resist the expression of A type endogenous genes. Sepals of the over expression vector transgenic petunia was erect, the corolla tube diminish dramatically and the petal tip was wrinkled like. Color of the anti-sense vector transgenic petal faded and the transgenic tobaccos were unopened. Interference vector transgenic plants were still small and subject to further observation. |
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