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为了解四川地区结瘤大豆根际土壤是否存在与紫云英、苜蓿和三叶草共生结瘤的根瘤菌及其多样性,从四川盆地采集不同种植模式下结瘤大豆根际土壤样品31份,利用紫云英、苜蓿和三叶草进行盆栽捕获试验以获得共生根瘤,从根瘤中分离纯化出根瘤菌,对其进行表型和分子鉴定;并将根瘤菌回接到紫云英、苜蓿、三叶草和大豆根际以检测根瘤菌的寄主范围.在捕获实验中,苜蓿和三叶草分别在6个土壤样品中共生结瘤,而紫云英只在2个土样中共生结瘤,并从共生根瘤中分离出14株根瘤菌;16S r DNA基因序列的相似性分析表明这14株根瘤菌均属于根瘤菌属(Rhizobium),且16S r DNA基因的系统发育树揭示它们分属于根瘤菌属的不同种;这14株根瘤菌在回接实验中都只能让捕获豆科植物结瘤,而不能让其他3种豆科植物结瘤.本研究结果表明,四川地区结瘤大豆根际土壤中的紫云英根瘤菌(Rhizobium astragali)、苜蓿根瘤菌(R.meliloti)和三叶草根瘤菌(R.leguminosarum var.trifolii)资源较少,其与大豆根瘤菌(Bradyrhizobium japonicum)分属于不同的属,且根瘤菌的回接实验进一步证明了根瘤菌的寄主专一性.
In order to understand the existence and diversity of rhizobia co-occurring in the rhizosphere soil of nodulated soybean in Sichuan Province, 31 nodulated rhizospheric soils were collected from different habitats in Sichuan Basin. Chinese cabbage, alfalfa and clover were harvested to obtain symbiotic nodules. Rhizobium was isolated and purified from nodules and phenotypically and molecularly identified. The rhizobia was ligated back to the plants of Astragalus sinicus, alfalfa, clover and soybean In the rhizosphere, the host range of Rhizobia was detected.In the trapping experiments, alfalfa and clover symbiotic nodulation in six soil samples, while the milk vetch nodulated only in two soil samples and isolated from symbiotic nodules A total of 14 rhizobia strains were obtained. The similarity analysis of 16S r DNA sequence showed that these 14 Rhizobium belonged to Rhizobium, and the phylogenetic tree of 16S r DNA revealed that they belonged to different Rhizobium species. All 14 strains of Rhizobium were able to catch nodules of leguminous plants only in the relay-back experiment, but could not nodulate the other three leguminous plants.The results of this study showed that the purple cloud in rhizosphere soil of nodulated soybean in Sichuan Province English root nodules Rhizobium astragali, R. meliloti and R. leguminosarum var. Trifolii have less resources and belong to different genera with Bradyrhizobium japonicum, The experiment further confirmed the rhizobial host specificity.