Determination Of Stress Tolerance And Selected Cross Breeding Of The Progeny Of The Ground Cover Chrysanthemum 'Fall Color' Integrated DREB1A Gene

Ground cover Chrysanthemum plays an important role in landscape in the world.This paper is about transgenic ground cover Chrysanthemum 'Fall Color'(Chrysanthemum grandiflora(Ramat.)Kitamura 'Fall Color') introduced AtDREB1A gene,a stress-inducible transcription factor driven by 35S cauliflower mosaic virus(CaMV) promoter or stress inducible rd29A,and with the tolerance of low temperature,drought and saline,an excellent cultivar of Chrysanthemum.Experiment material was propagated by tissue culture. Determination of low temperature stress and high temperature stress of transgenic progeny was tested.Cross breeding technology was used between transgenic strains and other excellent ground cover Chrysanthemum cultivars.Then test ornamentals and exogenous gene through molecular detection of filial generations.The purposes of these experiments are to obtain middle material of new ground cover Chrysanthemum cultivars with great phenotype and stress tolerance,and to provide theoretic basis and actual basis for combination of gene engineering breeding and conventional cross breeding of ground cover Chrysanthemum.Transgenic strains were propagated by tissue culture,lateral meristem as explants.The survival rate of transplants is high in wintering open field,and the transplants grow normally. Compared with the wild type,the height,crown width,branch angels of the transgenic strains present phenotypic difference,while their flower period and flower color don't present phenotypic differences.In 35℃high temperature conditions,compared with the wild type,the heat tolerance of rd29A:DREB1A transgenic strains increase and 35S:DREB1A transgenic strains further increase.There is a relationship between cell membrane injury rate,malondialdehyde content, soluble protein content and the expression of integrated exogenous gene.We speculate that the introduction of exogenous gene DREB1A can affect the various metabolic regulations in high temperature conditions,thereby strengthen the capacity of cell membrane self-protection and membrane lipid peroxidation.And Chrysanthemum has an adaptation process to the heat stress The experiment of paraffin section observation about the bud differentiation process indicates that all strains bud differentiation is restrained in 38℃high temperature conditions Compared with the wild type,the restrained degree of transgenic strains is weaken.The above proves that capacity to adapt high temperature of transgenic strains is strong than the wild type.All confirms that the excess expression of the AtDREB1A gene from Arabidopsis can enhance ground cover Chrysanthemum heat tolerance.The experiment illustrates,once the bud differentiation,LD can not reverse the bud differentiation process.The temperature from November 2006 to January 2007 in Harbin:the average temperature is-10.0℃;the extremely low daily temperature is -23℃;the extremely low soil temperature is—18.7℃.Compared with the wild type,the survival rate of transgenic strains is high,and their capacity of overwintering in the open field increases.The cross breeding technology of non emasculation artificial pollination was used, transgenic plant integrated AtDREB1A gene as the female parents,the better ornamental and widely used cultivars such as 'DongLRX' as the male parents.Strains setting percentage and seed germination rate is high,filial generations grow and develop well.Exogenous gene tested by the PCR technology,proves Exogenous gene can be transferred to the by sexual cross breeding Many characters such as flower color,height and crown width present character segregation.22 filial generations with exogenous gene and some filial generations with excellent character have been selected from F1.