Arabidopsis Phya Gene Cloning And Transformation Of Chrysanthemum

Ground-cover Chrysanthemum is a group of Chrysanthemum morifolium,which is applied greatly in gardens and landscape due to its rich colours,faster growth rate and excellent ornamental value.Flowering time is an important character of chrysanthemum.It is easily affected by light and temperature.This greatly limited the further application of ground-cover chrysanthemum.In order to creat an early flowering and weak light tolerant chrysanthemum, the Arabidopsis phyA gene is transferred into the genome of Ground-cover Chrysanthemum 'Fenditan' by agrobacterium mediated transformation in our research.Phytochrome is one of the most important photomorphogenic photoreceptors.In Arabidopsis the family of Phytochrome photoreceptors is encoded by five genes,designated phyA to phyE.ThephyA gene,unlike the other pytochrome family members,has a high level of expression in darkness.Precious studies have already shown,phyA acts in promotion of flowering,overexpression ofphyA results in extreme dwarfism in mature plants.In our research,transformation system for 'fenditan' was established.In this system,leaf explants which were pre-cultured for 24 hours were the most suitable for transformation by Agrobacterium tumefaciens.The suitable concentration ofAgrobacterium tumefaciens and the suitable dipping time for transforming was OD600=0.4 and 10 minutes.Co-cultivation for 2 days at 24℃after infection,the explants were then put in delayed cultivation for 4 days.This is necessary for successful transformation.Then the explants were cultured in shoot induction medium with 5 mg/L kanamycin until the regenerated shoots developed.Once the regeneration shoot was 1cm long it was transformed into 1/2 MS for root induction.We have got 46 regenerated plants totally up to now.Only 4 of them were proved to be transgenic palnts witth PCR detection.One of the 4 phyA transgenic lines was created in 2008,named T1;the others were created in 2009.T1 line was studied further.Under natural light,compared with the control plants,the flowering time of T1 transgenic plants was 16 days and 22 dyas earlier respectively in 2008 and 2009.Petal color of T1 plants is yellow,while that of the control plant is pink.The leave of T1 plants was greener than that of the control.Under weak light,compared with the control, T1 plants grew with longer petiole,bigger and thinner leaves.