沙棘根瘤内生菌株库构建与微生物多样性分析

  目的  沙棘Hippophae rhamnoides根瘤中拥有丰富的微生物资源，探究根瘤内生菌微生物多样性，比较高通量测序与纯培养方法的优劣。  方法  运用16S rRNA高通量测序技术探究沙棘根瘤内生菌相对丰度和多样性差异，并对根瘤内生菌进行分离纯培养，以完成内生菌株库初步构建。  结果  ①传统分离方法得到纯培养菌株96株，共4门8纲8目13科19属。在门分类水平上，相对丰度较高的主要为变形菌门Proteobacteria和厚壁菌门Firmicutes。②高通量测序分析6组样品(M1~M6)中共有14门34纲89目148科314属。在门分类水平上，相对丰度较高的主要为放线菌门Actinobacteria和变形菌门，两者相对丰度之和为87.50%~97.10%。在属的分类水平上，弗兰克氏菌属Frankia占绝对优势，相对丰度为20.12%~74.81%，平均相对丰度为51.49%。③高通量测序与纯培养方法的结果有明显差异，在科和属的分类单元上差异较大。  结论  2种方法都体现沙棘根瘤内生菌的多样性，但纯培养方法仅能够分离到部分内生菌，难以评估物种组成及相对丰度。高通量测序分析能够更为全面地反映多样性信息，且为优化纯培养条件而分离特定物种奠定基础。图3表3参41

Construction of endophytic strain bank of seabuckthorn nodule and an analysis of microbial diversity

  Objective  This study is aimed to conduct an investigation of the microbial diversity endophytic bacteria in rhizobia of Hippophae rhamnoides which play an important role in nitrogen fixation and plant growth and research the construction of rhizobia endophytes.   Method  With the employment of 16S rRNA high-throughput sequencing technology, an exploration was conducted of the relative abundance and diversity of endophytes in Ulange wood rhizobia of Mongolian H. rhamnoides in Dengkou County before the endophytes were isolated and purely cultured to complete the construction of endophytic strains library.   Result  (1) A total of 96 pure culture strains were obtained by traditional isolation, with 4 phyla, 8 classes, 8 orders, 13 families, and 19 genera and Proteobacteria and Firmicutes are of relatively higher relative abundances in terms of phyla classification. (2) With an analysis of 6 groups of samples (M1?M6) consisting of 14 phyla, 34 classes, 89 orders, 148 families, and 314 genera by high-throughput sequencing, Actinomycota and Proteobacteria have higher relative abundance, and the sum of the relative abundance of the two is between 87.50%?97.10% in terms of phylum classification. As for the taxonomic level of the genus, the genus Frankia occupies an absolute advantage with the relative abundance being between 20.12% and 74.81%, and the average relative abundance being 51.49%. With M5 having the largest Shannon and Simpson indexes and M4 having the lowest species diversity, both M5 and M6 have higher levels of species abundance whereas M4 species has the lowest abundance. (3) The results of high-throughput sequencing and pure culture methods are significantly different, especially in the taxa of families and genera.   Conclusion  Both methods reflected the diversity of endophytes in sea buckthorn rhizobia, but the pure culture method could only isolate some endophytes, and it was difficult to evaluate the species composition and relative abundance. Also, high-throughput sequencing analysis could reflect the diversity better and lay the foundation for optimizing pure culture conditions for the isolation of specific species. Ch, 3 fig. 3 tab. 41 ref.]