滇朴丛芽病植原体的分子鉴定

Molecular identification of Celtis kunmingensis plexus bud phytoplasma

滇朴Celtis kunmingensis Cheng et Hong是云南的乡土树种,适宜全国大部分地区种植,极具观赏价值,是近年来最热门的绿化首选树种―绿化行道树,云南部分地区滇朴近年常表现丛芽的症状。本研究采用形态学与分子生物学结合的方法,对染病的幼嫩枝条进行扫描电镜(SEM)观察;利用16S rDNA植原体通用引物P1/P7和R16F2/R16R2进行常规PCR和巢式PCR,分别获得1.8 kb和1.2 kb的特异性基因片段,将该特异性片段与其他已知分类地位的植原体16S rDNA片段进行同源性比对分析,同时利用邻接法(NJ)构建系统发育树。结果表明在染病的滇朴韧皮部组织中可见植原体存在,滇朴丛芽病植原体与芝麻叶状植原体同源性高达99.40%,通过系统发育树可进一步推测滇朴丛芽病植原体是属于16SrⅠ-B亚组成员,本研究结果为该病害的诊断与防治提供了理论依据。

Celtis kunmingensis,a native tree species in Yunnan province,suitable for planting in most parts of China,and it has great ornamental value.In recent years,it is the most popular greening preferred tree species,named as the greening street tree.The diseases of plexus bud of C.kunmingensis was found in some parts of Yunnan province.In this study,morphological and molecular biological methods were used to observe the infected tender branch tissue under scanning electron microscopy(SEM).By direct PCR and nested PCR with 16S rDNA phytoplasma primers P1/P7 and R16F2/R16R2,the specific gene fragments of 1.8 kb and 1.2 kb were obtained,respectively.The 16S rDNA fragment of phytoplasma was compared with other phytoplasmas with known taxonomic status,and the phylogenetic tree was constructed based on Neighbor-Joining(NJ)method at the same time.The results showed that phytoplasmas were found in the phloem tissues of susceptible C.kunmingensis.The homology of C.kunmingensis plexus bud phytoplasma and sesame phyllody phytoplasma was as high as 99.40%.Through phylogenetic tree,we can further infer that phytoplasma is a 16Sr Ⅰ-B subgroup member,which provides theoretical basis for the diagnosis and control of the disease.