MONITORING OF RHIZOBLA ON THE ROOT SYSTEM AND ANALYSIS OF THE MECHANISM OF NODULATION IN SOME LEGUMES USING A GUS MARKED STRAIN

利用gus标记的根瘤菌对几种豆科植物根瘤菌侵染过程的多样性进行了研究。研究结果如下：1.为了获得gusA标记的慢生根瘤菌USDA 3024、B.sp. U9709-A和茎瘤固氮根瘤菌U9709-SRS菌株，用含有β-葡萄糖醛酸苷酶操纵子（pCAM 121上的mTn 5 SSgusA 21）的质粒与大肠杆菌S17-λ pir菌株进行接合实验。在X-Gluc缓冲液中培养时，各gus标记菌株形成的根瘤在Clotalaria、Arachis和田菁中均呈现蓝色. gus标记的菌株在各豆科植物中的固氮酶活性和固氮能力与亲本菌株没有差异.生长袋法适用于gus标记菌株在猪屎豆幼苗中的侵染试验.显微镜下观察发现，猪屎豆2日龄幼苗主根根毛中存在侵染线状结构.硝酸盐对花生根瘤数和固氮酶活性有抑制作用.长喙田菁茎瘤起源于子叶节部位，随后在茎表面形成.目前正在利用gus标记的菌株进一步研究根瘤菌感染过程的多样性以及各种环境条件对根瘤菌生长和固氮的影响。另一种标记基因GFP也将用于这些实验。

The diversity in the infection process of rhizobia to some legumes was investigated by using a gus-marked strain. The results are summarized as follows.1. To obtain gusA-marked strains of Bradyrhizobium sp. USDA3024, B.sp. U9709-A, and Azorhizobium caulinodans U9709-SRS, conjugation experiments were performed with the strain of E.coli S17-λ pir with plasmid containing the β-glucuronidase operon (mTn5SSgusA21 on pCAM121). Root nodules formed by each gus-marked strain developed blue color on incubation in X-Gluc buffer in Clotalaria, Arachis and Sesbania.2. The gus-marked strain did not differ from the parent strain in nodulation capability and nitrogenase activity in each legume.3. Growth pouch method was adequate for infection test of a gus-marked strain in Crotalaria seedling.4. With a microscopic observation, an infection thread-like structure was found in the root hairs of tap root in 2 day-old seedling of Crotalaria juncea.5. Number of root nodule and nitrogenase activity in peanut plants were inhibited by application of nitrate to culture solution.6. Stem nodules of Sesbania rostrata were initiated at the site of the cotyledonary node and followed by forming on the surface of stem.7. Further investigation of the diversity in the rhizobia-infection process and the effect of various environmental conditions on nodulation and nitrogen fixation is now in progress using the gus-marked strain. Another marker gene, GFP, would be also used for those experiments.