| Wheat is one of the most important food crops in China and the world, using transgenic technology for orientation improvement of wheat has attracted great attention and it will gradually become an effective assistance ways for developing new varieties with high-quality and high yield. Compared with rice, corn and other major food crops, wheat was difficult for plant regeneration with tissue culture, and its genetic transformation rate was very low. All this greatly retard the application of transgenic technology in cultivation of new wheat varieties. Therefore, improve regeneration and genetic transformation rate of wheat tissue culture is an important foundation for transgenic research.19 excellent winter wheat and 1 spring wheat cultivars in Huanghuai wheat area were used as test materials in the present study. One purpose of this study is to optimize and establish efficient regeneration system of mature embryos adapted to a variety genotypes of wheat. The second is to selected wheat genotypes with high regeneration frequency from the varieties investigated. Finally, genetic transformation was carried out in the wheat varietie with high regeneration rate by gene gun-mediated to obtain transgenic plants. The aim of this study was to lay a sound basis for transgenic wheat research in Huanghuai area and Shaanxi Province. The main results obtained were as follows:1. Optimization of regeneration system of wheat tissue culture from mature embryo Callus induction frequency of whole mature embryo with half longitudinal cut was obviously greater than half embryo by crosscut, half embryo by longitudinal cut or mashed embryo, moreover, the quality of callus formation from mature embryo with half longitudinal cut was best. Among six kinds differentiation media with different hormone ratio,3 differentiation medias with hormone matching as the follows,i.e. B. IAA 1.0 mg/L, ZT 2mg/L; E. ZT 10mg/L, 2, 4-D 0.1 mg/L; F. 6-BA 1mg/L, NAA 0.3 mg/L, KT 4mg/L were suitable for mature embryo regenerating of all the three wheat genotypes,and the regeneration rate was much higher than those in the other three differentiation medias. 1/2 MS culture medium without auxin were much better to prompt to generatation of roots in the regenerated buds .2. Screening of genotypes with high regeneration rateTwelve of the 20 wheat cultivars investigated, had the regeneration rate more than 20%, among which the regeneration rate of Xinong 928, Ningchun 16 and Xinong 889 were as high as 52.7%, 77.6%, 36.9 %, respectively. Differentiation medium B is adaptable for callus culture of most genotypes. These wheat cultivars can be used as the receptor varieties in wheat transgenic research in this region.3. Preliminary study of wheat transformation by biolistic-mediatedXinong928 was successfully transformated by the microprojectile bombardment, and 1200 callus be transformated. we gained 28 regeneration plants, and four PCR positive plants with the target genes Zm-R-S, transformation rate was 0.33 percent, and 14 positive plants with the screening Bar marker genes, transformation rate was 1.2 percent. Results indicated that 6 cm target distance was adapter for a higher regeneration rate of mature embryo of Xinong 928.The mature embryos of Xinong 928 was the best receptor materials for transformation. |
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