Study On Enhancing Boron Efficiency In Brassica Napus Via Overexpression Of BOR1

Boron (B), one of the necessary micronutrients for plant growth, is absorbed and utilized by plants in the form of H3BO3 in soils. In China, especially in the middle and down region of Yangtze River, soil available B deficiency is more severe and the application of B fertilizer has become an essential agronomic measure in B-sensitive crops production, such as Brassica napus and cotton. It was reported that overexpression of B transporter BOR1 in Arabidopsis increased boron transportation and improved seed yield under B-limiting condition.In this research, we transformed BOR1 into Brassica napus cultivar "Weatar 10" and obtained transgenic plants overexpressing BOR1. Then different phenotype identification experiments were carried out to study growth and B efficiency of transgenic plants under low B conditions. The main results were as follows:1. Generation and screening of transgenic plants.13 transforments overexpressing BOR1 gene were obtained By Agrobacterium mediated method. After self-breeding, We got 3,3 and 5 non-segrateing family lines from three transgeneic T2 generation lines, L1, L2, L3, respectively, by Kan screening and PCR identification. Further, we used seeds of the T3 generations to verify that, and seeds of T4 generation ware harvsted.2. Expression of extraneous gene BOR1 and copies identification. Analyzing the expression of BOR1 in different growth stages via RT-PCR showed that transgenic plants could stablely express BOR1 on RNA level. The result from Southern blot analysis indicated that L1 and L2 were double-copys lines, and L3 was a single-copy line.3. Phenotype identification of transgenic plants in seedling stage. Results of hydroponic culture indicated that under severe low-B condition (0.15μM), growth of transgenic plants had advantage over WT, boron concentration and accumulation in shoot of 3 transgenic lines were enhanced to different extent compared with WT. Thereinto, shoot boron concentration of L1 and L2 were almost doubled higher than WT, and shoot boron accumulation of L1, L2 and L3 were 2-3 times higher than WT.4. Phenotype identification of T1 and T3 transgenic plants for whole-growing period. Results of pot trials were as follows:Results of T1 indicated that under moderate B-shortage condition (0.25 mg/kg), silique, seed yield and oil content of L1 and L2 were all increased, and seed yield of L1 is 43% higher than WT. For L3, shoot boron concentration under severe B-shortage condition (0.09 mg/kg) and seed boron concentration under moderate B-shortage condition (0.25 mg/kg) were increased obviously, while seed yield had no difference from WT. Results of T3 indicated that under different boron deficient conditions, shoot yield, plant height, available siliques, available branches and seed yield of L1 and L2 were improved compared with WT. Thereinto, shoot dry weight of L2 in bud stage was 15.8% higher than WT under moderate B-shortage condition (0.25 mg/kg). Seed yield of L1 and L2 were higher than WT under severe B-shortage condition (0.09 mg/kg), and increased by 3% and 10%, respectively, under moderate B-shortage condition (0.25 mg/kg). Results of the experiments indicated that transgenic plants can grow and set seed better than WT when suffering from boron deficiency.