Preliminary Study Of The Agrobacterium Tumefaciens-Mediated Genetic Transformation Of Soybean Cotyledonary Node

Soybean [Glycine max (L.) Merr.] originated in China, which is a diploid (2n=40) plant belong to Leguminosae Papilionoideae. It is the major grain and oil crop in the world. Soybean quality and yield are affected by various of stresses, like diseases, insects, and weeds. The conventional breeding takes up a long time period, with limited genetic resources and slow efficiency. So researchers are trying to use transgenic methods to improve soybean yield, quality and resistance to stresses.Agrobacterium-mediated transformation using soybean cotyledonary node is the most mature way, but soybean transformation is more difficult than other crops because of the low transformation ratio and poor reproducibility. The genotype of the acceptor material is one of the most important factors in the soybean transformation. So we made researches on the genotypes of the acceptor material and the culture conditions to establish a suitable transformation system. The main results were as follows:In this study, we use Agrobacterium-mediated cotyledonary node transformation system to transform the soybean varieties including Nannong86-4, Nannong88-31, Nannong88-1, Nannong99-6, Houzimao, Dalihuang, Jack, and Williams82et al. Firstly, the genotypes of receptors were studied to determine the sterilization times, available rates and the growth potentials for different varieties after germination and made a comparison of shoot regeneration frequency among different varieties. The results showed that Jack required the longest time, between6.5h-7h, Houzimao taked the shortest time, about4.5h-5h, and others were mainly between5h-6h. Nannong88-1, Jack and Houzimao had higher availability than Nannong88-31. Houzimao had the highest regeneration frequency, up to82.7%, and the regeneration ratio of Nannong86-4was the lowest, only39.4%.Secondly, we made some optimization on the culture conditions of Agrobacterium-mediated soybean cotyledonary node, including the impacts of different screening concentrations on bud induction and shoot elongation, and the technics of rooting and seedlings transplanting. The results indicated that under3-5mg/L Hyg-B screen, the budding rates and the proliferation numbers were decreased with the increasing of the screen concentration. In the process of shoot elongation, Nannong88-1and Dalihuang performed better with the highest stem-drawing rate,56.5%and52.6%respectively. The tolerance to Hyg-B from high to low was Jack, Nannong88-1, Dalihuang, and Houzimao. In addition, we found that B5culture-mediated with NAA was more profitable to the growth and rooting of the seedlings than1/2B5culture-mediated with IBA.