The Optimization Of Wheat Culture System And Young Spike Transformation System

Wheat is one of three major food crops around the world, imporving the yield of wheatis badly in need of solution to ensure food supply, as the growing problem of population andfood crisis. With many years’ domestication and improvement, it’s difficult to increaseproduction only by traditional breeding techniques. Thus, the applying of transgenictechnology plays more significant role in genetic breeding and gene function studies.In the study, an optimized wheat tissue culture system of mature embryos and young spikeswere set up. Meanwhile, particle bombardment and agrobacterium-mediated transformationsystem of young spikes were also set up. With these two methods, the EhHOG gene wassucceeded in importing into5different genotypes, results are as follows:7genotypes were utilized to optimize the mature embryo tissue culture system, concludingthe impacts of diverse kinds of induction medium, way of inoculation and pretreatments.Results indicate that callus rate varies little among different genotypes, reaching90%, whilethe embryogenic callus rate and differentiation rate varies between10.72%and34.72%;Soaking seeds in4℃for15h, then inoculation by cut is more likely to achieve higher callusrate and differentiation rate; The callus rates and embryogenic callus rates of Xinong509,induced by8kinds of induction medium, show significant differences. The selected Mginduction medium could obtain the highest callus rate (93.37%) and differentiation rate(13.22%).196commercial varieties or strains were examined by the mentioned regenerationsystem, obtaining10varieties or strains, whose callus rates varies between95.90%and100%,holding higher callus rates than control. Othervise, the10varieties (strains) also possesshigher differentiation rate than control, with the highest strains “D bai C23” reaching36.78%.All the selected genotypes are easier to apply in genetic improvement, because of their betteragronomic traits and yield ability.The study focuses on factors, which has a significant impacts on the tissue culture ofyoung spike, like sampling date, length of spike and different parts of spikes. Results showthat, compared with0.5and2.5cm, the spikes, length of1.0cm, could induced shortest timeof callusing at50%, highest callus rates and differentiation rates. There is no remarkabledifferences of plant regeneration between0.4and1.0cm, but the utilization rate of0.4cm is obviously higher than1.0cm. Referring to the effects of different parts of young spikes, theplant regeneration rates of proximal and middle parts of young spikes is higher than the top.The plant regeneration rates of young spike and mature embryo were nearly the same,however, the differentiation rates were just the other way around. The comparision betweenplant regeneration rates of young spike and mature embryo revealed that, genotypes had lessimpacts on the culture of young spike than mature embryo, so optimizing the regenerationsystem by the selection genotypes with better agronomic traits would be more efficient thanother ways. And young spikes is just the oppsite.As is known to all, the overgrowth of agrobacterium has an effects on transformationrates. The study focuses on factors, like concentration of agrobacterium, co-culturetemperature and way of co-culture. The results reveals that the overgrowth being serious asthe rose of concentration of agrobacterium. In the mean time, there was no significant effectsof way of co-culture on agrobacterium overgrowth. Eventually,6positive regeneration plants,belonging to Xinong509and Zhengmai366, were obtained, and the positive rates are0.23%and1.07%in turn.The impacts of diverse kinds of high osmotic medium and bombardment pressure are asfollows: Compared with1100psi, bombarded with650psi could achieve higher GUStransient expression rate, but the receptor cells would also suffer more severe damages,resulting in relatively lower differentiation rate. With proper osmotic treatment, receptor cellscould increase their ablity of withstanding the bombardment damages. It is sufficient tomention here, the level of withstanding varied among different kinds of penetrating agent.That is, the GUS transient expression rate of sucrose and mannite were higher than maltoseand control. With the optimized transformation system,16positive plants among2766regeneration plants, belonging to16different grops, were obtained.7of16were treated with650psi bombardment pressure, while9of16were treated with1100psi.