Establishment Of Genetic Transformation System Of Mature Embryos Of Common Wheat By Agrobacterium And Preliminary Exploration Of Improving Conversion Rates

Wheat is one of the most important food crops in China and the world, using transgenictechnology for orientation improvement of wheat has attracted great attention and it willgradually become an effective assistance ways for developing new varieties with high-qualityand high yield. Compared with rice, corn and other major food crops, wheat was difficult forplant regeneration with tissue culture, and its genetic transformation rate was very low. Allthis greatly retard the application of transgenic technology in cultivation of new wheatvarieties. Therefore, improve regeneration and genetic transformation rate of wheat tissueculture is an important foundation for transgenic research.An effective wheat callus induction system via inducing embryonic callus, was set up through Wheat variety Shaanmai159mature embryos as exophyte. The results showed that the seeds were thoroughly disinfected and got less damage after dipping into sodium hypochlorite for20min, then5min in mercury bichloride. It was easy to strip the mature embryo and get better callus when the dipping time was between18h to20h. Better callus was got on MS medium than on1/2MS medium, when adding4mg/L2,4-D into MS medium, the callus would be better, meanwhile the inducing efficency was the highest. Compared to84.167%with complete embryos the ratio of embryonic callus induction was up to94.167%by using broken embryos. So the system was that first disinfecting seeds with sodium hypochlorite for20minutes, then mercury bichloride for5minutes, third dipping the seeds into water18h to20h for stripping embryos, finally breaking the embryos on MS medium with2,4-D(4mg/L),30g sucrose and7g agar.The HOG1gene was transferred into the shoot apical meristem of wheat with different genotypes via agrobacterium-mediated transformation to study its genetic transformation. Our results showed that when using the shoot apical meristem as receptors, genotypes could affect other than control the agrobacterium-mediated transformation system. The transformation frequency of To generation was influenced by different transformation methods, meanwhile aseptic condition and more agrobacterium transformation times could increase the To generation transformation frequency. We got1037seedlings after transformation, and186plants survived after selecting with kanamycin and contained the HOG1gene which were detected by PCR. So the To transformation frequency reached17.94%on average. Among the T1generation, detected by PCR and RT-PCR, only2seedlings with stable inheritance remained after excluding of wheat chimera, the transformation frequency was0.19%.