赞皇大枣枣疯病植原体分子分类

doi:10.3864/j.issn.0578-1752.2011.21.011

摘要/Abstract

摘要： 【目的】枣疯病是一种重要的植原体病害，本试验旨在明确惟一已知的天然三倍体品种--赞皇大枣枣疯病植原体的分类地位，为赞皇大枣枣疯病植原体分类研究提供参考。【方法】利用植原体16S rDNA通用引物对赞皇大枣枣疯病病株的DNA进行PCR扩增和克隆测序，通过BLAST比对进行序列分析，利用临位相连法构建16S rDNA系统演化树，并应用DNAstar软件进行虚拟RFLP分析。【结果】获得的赞皇大枣枣疯病植原体16S rDNA序列长度为1 850 bp（GenBank登录号GU184180），包括1 529 bp的16S rDNA基因全序列、264 bp的邻近间隔区序列及57 bp的23S rDNA部分基因序列。同源分析表明，赞皇大枣枣疯病病原16S rDNA序列与其它植原体的相似性在87%—98%，其中与榆树黄化 Elm yellows（EY）(GenBank登录号l33763)最高相似性为98%；与大多数二倍体枣品种的植原体序列相似率达99%以上；其虚拟RFLP图谱与4个16SrⅤ亚组的代表序列图谱差异显著,赞皇大枣枣疯病植原体延伸因子tuf基因的序列(GenBank登录号JN001985) 比对结果表明，与其它亚组序列同源性为52.3%—76.7%。【结论】三倍体赞皇大枣枣疯病病原属于榆树黄化组(16Sr V)，并与其中的B亚组亲缘关系最近；与二倍体品种的病原分类地位一致，说明枣疯病植原体在系统进化上比较保守。

关键词: 赞皇大枣, 枣疯病植原体, 16S rDNA, 分子分类

Abstract: 【Objective】 Jujube Witches’ Broom (JWB) is a serious disease caused by phytoplasma. In this paper, the taxonomic status of JWB-associated phytoplasma from Ziziphus jujuba Mill. ‘Zanhuangdazao’，which is the only natural triploid cultivar in Chinese jujube, was identified.【Method】16S rDNA gene was amplified by using specific conservative primers and the nested-PCR primers, then the PCR product was cloned and sequenced. Homology analysis, dendrogram and virtual RFLP were also obtained by using related software. 【Result】 One set of the conserved primers was designed to amplify a 1 850 bp fragment (GenBank GU184180) which included 16S rDNA (1 529 bp), adjacent spacer sequence (264 bp) and partial sequence of 23S rDNA (57 bp) by PCR from total DNA extracted from diseased Ziziphus jujuba Mill. ‘Zanhuangdazao’. Nucleotide sequencing of the amplified fragment indicated that JWB-associated phytoplasma from Ziziphus jujuba Mill. ‘Zanhuangdazao’ shared nucleotide similarity about 87% to 98% with other phytoplasmas from data of GenBank, and a maximum similarity at 98% with Elm yellows（EY）phytoplasma. A phylogenetic tree based on 16S ribosomal DNA sequences was constructed. The result of virtual RFLP showed a significant difference among JWB-associated phytoplasma from Ziziphus jujuba Mill. ‘Zanhuangdazao’ and 4 subgroups. The blast result of tuf also showed that JWB-associated phytoplasma from Ziziphus jujuba Mill. ‘Zanhuangdazao’ (JN001985) shared nucleotide similarity about 52.3% to 76.7% with 4 subgroups. 【Conclusion】 JWB-associated phytoplasma from Ziziphus jujuba Mill. ‘Zanhuangdazao’ was clustered into the elm yellows group (16Sr V) phytoplasma, and it shared the highest similarity with the B subgroups. Its taxonomic status is consisting with those diploid cultivars.

Key words: Ziziphus jujuba Mill.‘Zanhuangdazao’, JWB-associated phytoplasma, 16S rDNA, molecular classification

杨海旭, 王洋, 赵彦檩, 赵锦, 刘孟军. 赞皇大枣枣疯病植原体分子分类[J]. 中国农业科学, 2011, 44(21): 4429-4437.

YANG Hai-Xu, WANG Yang, ZHAO Yan-Lin, ZHAO Jin, LIU Meng-Jun. Molecular Classification of Jujube Witches’ Broom (JWB) Associated Phytoplasma from Ziziphus jujuba Mill. ‘Zanhuangdazao’[J]. Scientia Agricultura Sinica, 2011, 44(21): 4429-4437.

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