| Rice(Oryza Sativa L.) is one of staple food, and its planting area is accounted for 30% of the countrywide planting area. The brown planthopper(BPH) has become the most destructive insect pest of rice with the characterization of short growth cycle, rapid propagation, long-distance migration and explosiveness. For a long time, utilization of large-scale chemical drugs results consequences of increased cost, BPH variation, resistance enhancement, rampant population, environmental destruction. In addition, cycle of traditional breeding for pest resistance is long and anti-gene is limited. Resistance of varieties will gradually lose along with production and application. This leads to the result of few resistance varieties of high levels. Discovery of new BPH resistance genes and breeding for pest resistance have become important goal of rice breeding to control BPH propagation and damage. Mutant library is important resource for studying gene function in genome-wide level and T-DNA insertion mutants are important materials for new varieties cultivation and functional genomics research of rice. Thus, in research of new BPH resistance genes and resistance breeding, mutant materials have always attracted universal attention.In this study, rice mutant materials were preliminarily selected, using the methods of resistance identification on heading stage and mass seeding, and agronomic traits were investigated to provide clues for discovery and applications of BPH resistance genes. The main results of this study were as follows: 1. more than 80 parts of T-DNA insertion mutant materials were preliminarily evaluated to determine levels of resistance on heading stage. We selected four representative mutants, and their numbers were as follows: 1663, 705, 406, 790. These materials were identified as the third grade of the BPH resistance. 2. Agronomic traits were investigated on these four mutant materials. Except for mutant 705, plant height, spike length and effective panicles of the other three mutants were significantly lower than Zhonghua 11, and thousand kernel weight of four mutants were significantly lower than Zhonghua 11. These results suggested that changes of agronomic traits were related to the resistance of mutants. 3. Using the method of mass seeding identification, we found two resistance mutants. Their numbers were as follows: 1663, 705, and these materials were identified as the third grade of the BPH resistance. But mutant 406 and mutant 790 were susceptible to BPH on seedling stage. There were differences with result of identification on heading stage in field.Brachypodium distachyon is a new model system of gramineous plants with the characterization of small genomes, effective transformation system, self fertilization, and easy to cultivation. And B. distachyon has a strong total linear relationship with genomics of wheat. With the completion of diploid B. distachyon genome sequencing, B. distachyon has always attracted common concern. Establishment of an efficient genetic transformation system is very critical for study of B. distachyon gene function. Because of its easy operation, high conversion rate, low integrated copy and genetic stability, Agrobacterium-mediated transformation method is widely used in rice, wheat and canola, and transformation system of these crops have been matured. The transformation system of B. distachyon is still not thorough, and domestic researchs have not been consistent.In this paper, the explants were immature embryos of B. distachyon diploid ecotype Bd21, and callus were transformed by the binary vector pCAMBIA1301 containing the reporter gene GUS. Agrobacterium-mediated transformation system of B. distachyon was established and a number of transgenic lines were got. The main results of this study were as follows: 1. Construction of transformation system of B. distachyon. Proper size of immature embryos was between 0.3mm and 0.7mm, optimal concentration of 2,4-D was 2.5mg/L and callus culture time was no more than 7 weeks. The ratio of callus inducement was 86%. Proper concentration of Agrobacterium and suspension time were OD600= 0.6 and 5min, co-culture time was 3d and a higher conversion rate was got. Adding 40mg/L hygromycin was in favor of resistant callus screening, and the seedling regeneration rate was up to 6.5%. 2. Detection of resistance plants. Callus and regeneration seedlings showed blue after GUS staining. We extracted DNA of regenerated plants and amplified fragment of GUS gene. Result of PCR analysis showed GUS gene was integrated into the genome of B. distachyon. |
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