ANALYSIS OF THE MECHANISMS OF NITROGEN INHIBITION ON RHIZOBIA INFECTION ON PEANUT USING AGROBACTERIUM RHIZOGENES-MEDIATED TRANSGENIC HAIRY ROOTS

利用发根农杆菌介导法在花生成熟干种子上诱导出毛状根。与对照相比，转化根和未转化枝组成的复合植株具有丰富的根系质量、更多的侧枝和较高的分维。将慢生根瘤菌接种到复合植株上，诱导出转化根瘤。通过C_2H_2还原实验发现，转化后的花生根瘤在细菌区产生了豆血红蛋白，C_2H_2还原实验显示了固氮酶活性，并表现出根瘤皮质区扩大和从根瘤皮层向新生根形成等形态变化。植株干重和全氮含量因培养液补氮时间的不同而不同，即在生长早期补充氮素可增加根瘤数、根瘤鲜重和乙炔还原活性。利用该复合植物和该培养方法，进一步研究了硝酸盐抑制花生根瘤菌侵染的机理。

Agrobacterium rhizogenes-mediated transformed hairy roots were induced on dry mature seed of peanut (Arachis hypogaea) cv.Java 13. Composite plants consisting of transformed roots with non-transformed shoots showed a root system with abundant root mass, more lateral branching and high fractal dimension compared to the control plant. The inoculation of Bradyrhizobium sp.to the composite plants led to the induction of transformed root nodules. Those transformed root nodules showed production of leghemoglobin in the bacterial zone and nitrogenase activity by C_2H_2 reduction assay, and exhibited morphological alterations such as enlargement of nodule cortex region and de novo root formation from nodule cortex.The effect of nitrate nitrogen at levels of 300 ppm N was investigated on the growth, root nodule formation and nitrogenase activity in peanut (cv.Chiba-handachi) inoculated with Bradyrhizobium and cultured in solution medium. Dry weight and total nitrogen content of the plant differed in response to the duration of supplement of nitrogen to the culture solution, i.e.supplement at early growing stage increased nodule numbers, nodule fresh weight and acetylene reduction activity. Pegging and pod formation were found on this hydroponic culture system.Further investigation of nitrate inhibition mechanisms on rhizobia infection on peanut is now in progress using the composite plant and this culture method.