Agrobacterium-mediated Haloduric Gene HaBADH Transformation In Embryo Tip Of Soybean

Salination is one of main environmental factors that affect yield and quality of soybean (Giycine max L.),the accumulation of betaine in plants to the benefit of enhancing salt tolerance and drought resistance,and betaine aldehyde dehydrogenase is the key enzyme in the synthetic pathway of betaine.In the paper,on the basic of optimizing embryonic tips regeneration system of soybean and confirming the screening concentration, BADH gene which from Helianthus annuus was transformed by Agrobacterium-mediate in soybean,and the effects of genotypes of soybean strains and inoculate time of Agrobacterium on transformation was discussed, the experimental results was followingThe effect of11genotypes(JiLin30,JiLin35,JiLin47,JiYu91,HeiNong35, HeiNong37,HeiNong44,HeFeng35,SuiNong28,BeiJingxiaolidou,DongNong50)on adventitious buds from embryo tip was studied in soybean.The experiment results showed that JiLin35,HeiNong44and HeFeng35could use for transformation of soybean, because they had a better effect on inducing adventitious buds from embryo tip of soybean,and got a rate of adventitious buds beyond92.00%.In order to evaluate the tolerance of JiLin35,HeiNong44and HeFeng35to three screening reagents, embryo tips were culrued, in medium plus containing different concentrations of NaCl, kanamycinand glufosinate.The results showed that three genotypes had different tolerance to the three antibiotics,the appropriate concentration of NaCl to screen JiLin35,HeiNong44,HeFeng35was75,25and100mM respectively; the appropriate concentration of kanamycin to screen JiLin35,HeiNong44,HeFeng35was100mg/L;the appropriate concentration of glufosinate to screen JiLin35,HeiNong44,HeFeng35was0.2,0.2and0.6mg/L respectively.Three Agrobacterium strains (LBA4404,EHA105and C58) to which were used to inoculate JiLin35,HeiNong44and HeFeng35with pCAMBIA2300-HaBADH, then the three genotypes were inoculated with the more insensitive strain by different inoculate time.Embryo tips which were inoculated by three Agrobacterium strains were screened by NaCl or glufosinate.and then were induced rooting,and the resistant plants was detected by PCR.C58was fit to inoculate JiLin35, HeiNong44and HeFeng35;the appropriate inoculate time of Agobacterium to JiLin35,HeiNong44and HeFeng35was15-25min,30min and5-30min respectively.37positive transgenic plants of soybean were got after transformed JiLin35,HeiNong44and HeFeng35by optimal system of genetic transformation,and found that there was difference existed in PCR positive rate.