Embryonic Callus Induction And Genetic Transformation Of Basic Study In Rosa Chinesis’yueyuehong’

In order to establish a somatic embryogenesis and plant regeneration system for rose, the crucial effects of light quailities and plant growth regulators on somatic embryogenesis and plant regeneration in Rosa chinensis ’Yueyuehong’ were investigated. Besides, studies of antibiotic susceptibility testing, the convertion conditions and the transient expression rate of GUS in embryonic callus and bud antibiotic confrontational aspects of screening throngh using embryogenic callus as the genetic transformation receptors in regeneration system in Rosa chinensis ’Yueyuehong’ were observed. The main research contents and results were as follows:1.Establishment of somatic embryogenesis and regeneration in R. chinensis ’Yueyuehong’.Using the sterile tissue culture leaves of R. chinensis ’Yueyuehong’ as explant, TDZ on embryonic callus induction effects, different concerntrations of ABA and GA3on cell proliferation and different light qualities on the type of somatic embryogenesis and plantles regeneration frequency were explored.The results showed that a low concentration of TDZ favor induction of embryonic callus. Using TDZ alone was easy to induce embryonic callus ’fungus’ shape. TDZ in combination with2,4-D alone is better than the use of2,4-D alone. And SH+2,4-D3.0mg/L+TDZ0.5mg/L was good for callus induction. Embryogenic callus GA33.0mg/L and ABA1.0mg/L in combination conducive to the formation cotyledon embryo, the highest proliferation coefficient was3.75, and embryo growth in good condition. Therefor, SH+2,4-D1.0mg/L+TDZ0.1mg/L+GA33.0mg/L+ABA1.0mg/L was appropriate medium for embryonic callus proliferation and differentiation.Also, types of light conditions have significantly influence on somatic embryos growth, and different types of somatic embryogenesis have large differences in regeneration frequency. Shoot-like embryos without cotyledon(s) were mostly observed in dark condition with the regeneration frequency of22%. It is concluded that one or two cotyledons of normal somatic embryos were produced under red and white light conditions which with the regeneration frequency of45%and15%. Somatic embryos with thick and expanded cotyledon(s), which had potential regenerating capability and the highest proportion of normal embryos produced cotyledons, were mostly observed in red condition. 2. Established and optimized of embryogenic callus material for the receptor genetic transformation system in R. chinensis’Yueyuehong’.In order to establish and optimize the R. chinensis’Yueyuehong’genetic transformation system, embryogenic callus were used as the genetic transformation receptors. The results showed that Kanamycin has a certain sensitivity influence on embryonic callus. In the medium to add to the card75mg/L kanamycin inhibit embryonic callus growth.We have preliminarily determined an effevtive system for genetic transformation. The protocol was:OD6oo=0-6,25minutes for infection,3days for co-culture, and AS150μmol/L as co-culture medium. Largest number of resistant callus produced and the rate of GUS transient expression was58.35%. At this time a small amount of surrounding colonies of embryonic callus. Bud induction and resistance than the filter medium is suitable for the inhibition SH+2,4-D1.0mg/L+TDZ0.1mg/L+ABA1.0mg/L+GA33.0mg/L+Cef200mg/L. And obtained by PCR preliminary identification of’transgenic plants in R. chinensis ’Yueyuehong’.