›› 2005, Vol. 48 ›› Issue (5): 810-814.

• 研究论文 • 上一篇    下一篇

桃蚜自然种群初级和次级共生菌的分子鉴定

李正西,李定旭   

  1. 中国农业大学昆虫学系
  • 出版日期:2005-10-13 发布日期:2005-10-20
  • 通讯作者: 李正西

Molecular identification of the primary and secondary symbionts of the green peach aphid, Myzus persicae (Sulzer)

LI Zheng-Xi, LI Ding-Xu   

  1. China Agricultural University
  • Online:2005-10-13 Published:2005-10-20

摘要:

蚜虫不仅带有专性初级内共生菌,还常常带有不同类群的兼性次级共生菌。本研究采用真细菌16S rDNA的通用引物,从桃蚜Myzus persicae (Sulzer)烟草种群体内扩增出1.5 kb的共迁移目的条带,然后将其克隆,并在阳性克隆筛选时进行了RFLP分析。当采用EcoRⅠ进行单酶切分析时,目的片段被切割成~650 bp和~850 bp两个小片段;当采用EcoRⅠ和HindⅢ进行双酶切分析时,该蚜群共生菌酶切谱带被分成2组,其中1组(GroupⅠ)只具有一个EcoRⅠ酶切位点,而另1组(Group Ⅱ)不仅具有一个EcoRⅠ酶切位点,还具有一个HindⅢ酶切位点,而且GroupⅠ为优势共生菌群。在此基础上分别对2组共生菌的16S rDNA全序列(~1.5 kb)进行了测定,结果表明:GroupⅠ属于泛菌属Pantoea,与成团泛菌Pantoea agglomerans亲缘关系最近(同源性达99.70%),而Group Ⅱ属于蚜虫的专性内共生菌,即Buchnera aphidicola(同源性达99.50%)。这是在蚜虫体内存在泛菌次级共生菌的首次报道。
 

关键词: 桃蚜, 共生菌, Buchnera aphidicola, 泛菌, 分子鉴定, 16S rDNA

Abstract:

Aphids harbor not only primary endosymbionts, namely Buchnera aphidicola, but also different secondary symbionts. A single comigrating DNA band of about 1.5 kb was amplified from a population of the green peach aphid, Myzus persicae (Sulzer) collected from the experimental fields in China Agricultural University (Beijing) using a pair of “universal" primers designed based on the full 16S rDNA of a wide range of eubacteria. This target DNA was then ligated to T-vector and cloned into E. coli DH5α. The clones were analyzed using restriction fragment length polymorphism (RFLP). The results showed that EcoRⅠcut the insert into two sub-fragments (~650 bp and ~850 bp) in all assayed samples, while double digestion with EcoRⅠ and HindⅢ resulted in two different groups of RE (restriction enzyme) bands, with GroupⅠhaving only one EcoRⅠ site but Group Ⅱ having one EcoRⅠ site and one HindⅢ site. Meanwhile, GroupⅠis dominant. Based on RE grouping, the full 16S rDNA representative sequences from different groups were sequenced, and the phylogenetic analysis demonstrated that GroupⅠ belonged to Pantoea, closely related to P. agglomerans (homology: 99.70%), whereas GroupⅡwas closely related to Buchnera aphidicola, the obligate symbiont of aphids (homology: 99.50%). This is the first report that Pantoea exists in aphids.

 

Key words: Myzus persicae, symbionts, Buchnera aphidicola, Pantoea, molecular identification, 16S rDNA