含羞草、山蚂蟥等几种豆科植物根瘤菌的数值分类及16S rDNA PCR-RFLP分析

为揭示云南省豆科植物根瘤菌的资源多样性,采用数值分类、16S rDNA PCR-RFLP和16S rDNA全序列分析技术对云南省8个地区含羞草、山蚂蝗等几种不同豆科植物分离的48株根瘤菌菌株进行了遗传和表型多样性研究。16S rDNA PCR产物经HinfⅠ、MspⅠ、RsaⅠ和HaeⅢ4种内切酶消化后产生22种遗传图谱类型;数据经MVSP3.0软件聚类后,所有供试菌株在70%的相似性水平上分为4个分支,经16S rDNA全序列分析,确定分支Ⅰ为伯克霍尔德属(Burkholderia)类群,含1株菌株;分支Ⅱ为贪铜菌属(Cupriavidus)类群,含5株菌株;分支Ⅳ为根瘤菌属(Rhizobium)类群,包括29株菌株;而分支Ⅲ没有和参比菌株聚在一起,经测序为狭长平胞属(Stenotrophomonas)类群。通过对菌株的抗生素抗性、耐盐性、生长温度范围、BTB产酸产碱反应以及对3-酮基乳糖利用情况共计28个表型性状测定,结果显示所有菌株耐盐性较强,多数菌株可耐受60℃的高温。实验表明,分离自云南省的这些菌株具有丰富的遗传和表型多样性。

Numerical Taxonomy and 16S rDNA PCR-RFLP of Rhizobia of Some Legumes Isolated from Mimosa spp. and Desmodium spp.

In order to characterize the diversity and phylogeny of rhizobium in Yunnan of China,48 strains isolated from root nodules of Mimosa spp.and Desmodium spp.in 8 regions of Yunnan Province were studied with numerical taxonomy,16S rDNA PCR-RFLP and 16S rDNA gene sequencing analysis.23 genotype profiles were obtained from the fingerprinting of the 16S rDNA PCR-RFLP by four restriction endonucleases Hinf Ⅰ,Msp Ⅰ,Rsa Ⅰ and Hae Ⅲ.These strains were clustered into 4 branches at the similarity level of 70% by MVSP3.0 software.16S rDNA sequencing revealed that branch Ⅰ belonging to the genera of Burkholderia,include 1 strain;branch Ⅱ belonging to the genera of Cupriavidus,include 5 strains;branch Ⅳ belonging to the genera of Rhizobium,include 29 strains;branch Ⅲ belonging to the genera of Stenotrophomonas with no reference strains.The analysis of 28 phenotypic characteristic of these strains showed that most of these strains had a high level resistance to saline,some strains could grow under severe environment such as high content ration of antibiotics(300 μg/mL) and high temperature(60℃).All the results demonstrated a high genetic and phenotypic diversity of rhizobium nodulating legumes of Yunnan Province.