Gene Transformation And CDNA-SRAP Analysis In Rosa Chinensis Var. ’Yueyuehong’

A two-gene-pyramided (pBin438-MtDREB1C-nos-35S-RcXET) was constructed by researchers of our laboratory. The two genes were used in stress-tolerance improvement and germplasm innovation of rose. Transformed it into Rosa chinensis var.’Yueyuehong’ via Agrobacterium tumerfaciens mediation using axillary bud of stem. Compare with the no-transformed plants by cDNA-SRAP analysis. The main results and conclusions are summarized as follows:1. Established and optimized of axillary bud of stem for the receptor genetic transformation system in R. chinensis var.’Yueyuehong’.Using the axillary bud of stem of R. chinensis var.’Yueyuehong’ as explant,6-BA and GA3 on axillary bud induction effects, different concentrations of Cef and Kan on the growing of the buds and the shoot frequency were explored. The results showed that MS+6-BA 2.0mg/L+GA3 1.Omg/L was good for buds induction, the highest proliferation coefficient was 96.6%.In order to establish and optimize the R. chinensis var.’Yueyuehong’ genetic transformation system, axillary bud of stem were used as the genetic transformation receptors. Kanamycin has a certain sensitivity influence on axillary bud of stem. In the medium to add to the card 150mg/L kanamycin inhibit bud growth.We have preliminarily determined an effective system for genetic transformation. The resistant bud coefficient was 4.98%. Determination of physiological indexes showed that the transgenic plants enhanced the tolerance to low temperature.2. Established and optimized of the SRAP in R. chinensis var.’Yueyuehong’A reliable and effective PCR reaction system for detecting SRAP was developed. It was shown that SRAP technology is a useful molecular marker system for study the differences in gene expression. Each 30μl PCR reaction mixture consisted of 10×Taq Buffer with KC1 3μl; MgCl2 (25mM) 3.2μl. Samples were subjected to thermal profile for amplification in an oven thermocycler, the annealing temperature was increased to 50℃.Ten primer combinations selected from the total random primers, the polymorphic map showed that the differences in gene expression in no-transformed rose and the transformed rose.