花生慢生根瘤菌的多样性研究
摘要
本研究采用表型性状测定、16S rDNA PCR-RFLP、16S rRNA序列分析和16S-23SrDNA ITS PCR-RFLP分析等技术系统研究了从我国不同地域分离的55株花生慢生根瘤菌的遗传多样性及其在根瘤菌系统发育中的地位和相互关系。
表型聚类结果表明：在69％相似性水平上，所有供试菌株和参比菌株可以分为两大类群Ⅰ和Ⅱ。在75.4％的相似性水平上，群Ⅰ可分为ⅠA、ⅠB和ⅠC三个亚群。亚群ⅠA由来自重庆、襄樊、泰安的菌株共同组成；亚群ⅠB由来自襄樊和泰安的菌株组成；亚群ⅠC由来自襄樊的菌株组成。在78％的相似性水平上，群Ⅱ也可分为ⅡA、ⅡB和ⅡC三个亚群。亚群ⅡA是由来自重庆、襄樊和盐城的菌株组成；亚群ⅡB主要由来自许昌的菌株组成，还包括3株来自重庆的菌株；亚群ⅡC由6株参比菌株Bradyrhizobium japonicum USDA6、USDA110、USDA122和Bradyrhizobium elkanii USDA46、USDA76、USDA86组成。
16S rDNA PCR-RFLP和16S rRNA序列分析结果表明：在63％的相似性水平上，所有供试菌株与参比菌株可分为三大类群。群Ⅰ由TA15等5株供试花生慢生根瘤菌组成，该群菌株在系统发育上与B.japonicum和B.elkanii的参比菌株亲缘关系较远。群Ⅱ由参比菌株B.japonicum USDA6、USDA 110和USDA122等组成。群Ⅲ在相关系数为0.818时可以再分为亚群1和亚群2两个亚群，亚群1中包含TA2等42株菌株，亚群2由自盐城分离的YC10等8个菌株与参比菌株B.elkanii USDA46、USDA76和USDA86组成。
16S-23S rDNAITS PCR-RFLP分析在61％的相似性水平上，将所有供试菌株和参比菌株分为三大类群：其中群Ⅰ由TA15等5株供试花生慢生根瘤菌组成。群Ⅱ由参比菌株B.japonicum USDA6、USDA110、USDA122和B.elkanii USDA 46、USDA76和USDA86组成。群Ⅲ在相关系数为0.658时也可以再分为亚群1和亚群2两个亚群，亚群1由TA2等42株菌株组成，可以在相关系数为0.939时分为六个小群，；亚群2由来自江苏盐城的8个菌株组成，在相关系数为0.791时YC10与其他7个菌株分开，单独聚为一类。与16S rDNA PCR-RFLP相比，16S-23S rDNA ITS PCR-RFLP具有更高的解析度，供试菌株表现出更加丰富的遗传多样性。
Studies on diversity and phylogeny of slow-growing rhizobia isolated from Arachishypogaea in China was determined by analysis of phynotype, 16S rDNA PCR-RFLP, 16SrRNA gene sequence, 16S-23S rDNA ITS PCR-RFLP.
Results of phenotype test shown that all 55 rhizobia tested and 6 reference strainswere clustered into two groups(Ⅰ,Ⅱ) at the similarity of 69%. GroupⅠcould be dividedinto subgroupⅠA,ⅠB andⅠC. SubgroupⅠA contained rhizobia isolated from Chongqing,Xiangfan and Taian. SubgroupⅠB contained rhizobia isolated from Xiangfan and Taian.SubgroupⅠC contained rhizobia isolated from Xiangfan. GroupⅡalso could be dividedinto subgroupⅡA,ⅡB andⅡC. SubgroupⅡA contained rhizobia isolated fromChongqing, Xiangfan and Yancheng. SubgroupⅡB contained rhizobia isolated fromXuchang and Chongqing. SubgroupⅡC contained 6 reference strains B.japonicumUSDA110, USDA6, USDA122 and B. elkanii USDA76, USDA46 andUSDA86.
Results of 16S rDNA PCR-RFLP analysis and 16S rRNA gene sequence revealedthat all strains tested could be clustered into three groups at the similarity of 63%. GroupⅠcontained 5 slow-growing rhizobia including TA5, they were relatively far from B.japonicum and B. elkanii phylogenetically. GroupⅡconsisted of 3 reference strains ofB.japonicum USDA110, USDA6 and USDA122. GroupⅢcould be divided into twosubgroup. SubgroupⅠcontained 42 slow-growing rhizobia including TA2. SubgroupⅡcontained 8 slow-growing rhizobia isolated from Yancheng and the reference strains of B.elkanii USDA76, USDA46 and USDA86.
Results of 16S-23S rDNA ITS PCR-RFLP shown that strains tested could be dividedinto two groups at the similarity of 61%. GroupⅠcontained 5 slow-growing rhizobiaincluding TA5. GroupⅡcontained 6 reference strains B. japonicum USDA110, USDA6,USDA122 and B. elkanii USDA76, USDA46 and USDA86. GroupⅢcould be dividedinto two subgroup. SubgroupⅠcontained 42 slow-growing rhizobia including TA2 whichcould be subdivided into six groups at the similarity of 0.939. SubgroupⅡcontained 8slow-growing rhizobia isolated from Yancheng. At the similarity of 0.791, YC10 wasseparated from other 7 slow-growing rhizobia. Compared with 16S rDNA PCR-RFLP,ITS RFLP assay shown higher resolution.
表型聚类结果表明：在69％相似性水平上，所有供试菌株和参比菌株可以分为两大类群Ⅰ和Ⅱ。在75.4％的相似性水平上，群Ⅰ可分为ⅠA、ⅠB和ⅠC三个亚群。亚群ⅠA由来自重庆、襄樊、泰安的菌株共同组成；亚群ⅠB由来自襄樊和泰安的菌株组成；亚群ⅠC由来自襄樊的菌株组成。在78％的相似性水平上，群Ⅱ也可分为ⅡA、ⅡB和ⅡC三个亚群。亚群ⅡA是由来自重庆、襄樊和盐城的菌株组成；亚群ⅡB主要由来自许昌的菌株组成，还包括3株来自重庆的菌株；亚群ⅡC由6株参比菌株Bradyrhizobium japonicum USDA6、USDA110、USDA122和Bradyrhizobium elkanii USDA46、USDA76、USDA86组成。
16S rDNA PCR-RFLP和16S rRNA序列分析结果表明：在63％的相似性水平上，所有供试菌株与参比菌株可分为三大类群。群Ⅰ由TA15等5株供试花生慢生根瘤菌组成，该群菌株在系统发育上与B.japonicum和B.elkanii的参比菌株亲缘关系较远。群Ⅱ由参比菌株B.japonicum USDA6、USDA 110和USDA122等组成。群Ⅲ在相关系数为0.818时可以再分为亚群1和亚群2两个亚群，亚群1中包含TA2等42株菌株，亚群2由自盐城分离的YC10等8个菌株与参比菌株B.elkanii USDA46、USDA76和USDA86组成。
16S-23S rDNAITS PCR-RFLP分析在61％的相似性水平上，将所有供试菌株和参比菌株分为三大类群：其中群Ⅰ由TA15等5株供试花生慢生根瘤菌组成。群Ⅱ由参比菌株B.japonicum USDA6、USDA110、USDA122和B.elkanii USDA 46、USDA76和USDA86组成。群Ⅲ在相关系数为0.658时也可以再分为亚群1和亚群2两个亚群，亚群1由TA2等42株菌株组成，可以在相关系数为0.939时分为六个小群，；亚群2由来自江苏盐城的8个菌株组成，在相关系数为0.791时YC10与其他7个菌株分开，单独聚为一类。与16S rDNA PCR-RFLP相比，16S-23S rDNA ITS PCR-RFLP具有更高的解析度，供试菌株表现出更加丰富的遗传多样性。
Studies on diversity and phylogeny of slow-growing rhizobia isolated from Arachishypogaea in China was determined by analysis of phynotype, 16S rDNA PCR-RFLP, 16SrRNA gene sequence, 16S-23S rDNA ITS PCR-RFLP.
Results of phenotype test shown that all 55 rhizobia tested and 6 reference strainswere clustered into two groups(Ⅰ,Ⅱ) at the similarity of 69%. GroupⅠcould be dividedinto subgroupⅠA,ⅠB andⅠC. SubgroupⅠA contained rhizobia isolated from Chongqing,Xiangfan and Taian. SubgroupⅠB contained rhizobia isolated from Xiangfan and Taian.SubgroupⅠC contained rhizobia isolated from Xiangfan. GroupⅡalso could be dividedinto subgroupⅡA,ⅡB andⅡC. SubgroupⅡA contained rhizobia isolated fromChongqing, Xiangfan and Yancheng. SubgroupⅡB contained rhizobia isolated fromXuchang and Chongqing. SubgroupⅡC contained 6 reference strains B.japonicumUSDA110, USDA6, USDA122 and B. elkanii USDA76, USDA46 andUSDA86.
Results of 16S rDNA PCR-RFLP analysis and 16S rRNA gene sequence revealedthat all strains tested could be clustered into three groups at the similarity of 63%. GroupⅠcontained 5 slow-growing rhizobia including TA5, they were relatively far from B.japonicum and B. elkanii phylogenetically. GroupⅡconsisted of 3 reference strains ofB.japonicum USDA110, USDA6 and USDA122. GroupⅢcould be divided into twosubgroup. SubgroupⅠcontained 42 slow-growing rhizobia including TA2. SubgroupⅡcontained 8 slow-growing rhizobia isolated from Yancheng and the reference strains of B.elkanii USDA76, USDA46 and USDA86.
Results of 16S-23S rDNA ITS PCR-RFLP shown that strains tested could be dividedinto two groups at the similarity of 61%. GroupⅠcontained 5 slow-growing rhizobiaincluding TA5. GroupⅡcontained 6 reference strains B. japonicum USDA110, USDA6,USDA122 and B. elkanii USDA76, USDA46 and USDA86. GroupⅢcould be dividedinto two subgroup. SubgroupⅠcontained 42 slow-growing rhizobia including TA2 whichcould be subdivided into six groups at the similarity of 0.939. SubgroupⅡcontained 8slow-growing rhizobia isolated from Yancheng. At the similarity of 0.791, YC10 wasseparated from other 7 slow-growing rhizobia. Compared with 16S rDNA PCR-RFLP,ITS RFLP assay shown higher resolution.
引文
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