柑橘黄龙病病原菌非洲种和美洲种23S-5S rDNA序列的克隆及分析

doi:10.3864/j.issn.0578-1752.2011.17.022

中国农业科学 ›› 2011, Vol. 44 ›› Issue (17): 3683-3693.doi: 10.3864/j.issn.0578-1752.2011.17.022

柑橘黄龙病病原菌非洲种和美洲种23S-5S rDNA序列的克隆及分析

廖惠红, 李杨瑞, 杨丽涛, 徐宁

1.广西农业科学院园艺研究所
2.广西作物遗传改良生物技术重点开放实验室
3.广西大学亚热带农业生物资源保护与利用国家重点实验室

收稿日期:2010-10-25 修回日期:2011-01-19 出版日期:2011-09-01 发布日期:2011-02-28
通讯作者: 通信作者李杨瑞，E-mail：liyr@gxaas.net
作者简介:廖惠红，Tel：0771-3245047；E-mail：liaohuihong2001@gmail.com
基金资助:
国家自然科学基金项目（31160390）、广西农业科学院基本科研业务专项（201009Z（基））、广西科学研究与技术开发计划项目（桂科攻1123003-2A）、广西自然科学基金回国基金项目（2011GXNSFC018012）、广西壮族自治区人才小高地项目（桂发办[2004] 47号）、广西农业科学院博士启动基金项目

23S-5S rDNA Sequence Cloning and Analysis of Candidatus Liberibacter africanus and americanus of Citrus Huanglongbing

LIAO Hui-Hong, LI Yang-Rui, YANG Li-Tao, XU Ning

1.广西农业科学院园艺研究所
2.广西作物遗传改良生物技术重点开放实验室
3.广西大学亚热带农业生物资源保护与利用国家重点实验室

Received:2010-10-25 Revised:2011-01-19 Online:2011-09-01 Published:2011-02-28
Contact: Hui-Hong LIAO

摘要/Abstract

摘要： 【目的】克隆柑橘黄龙病病原菌非洲种和美洲种23S-5S rDNA序列并和亚洲种相应区域进行比对，阐明黄龙病3个种核糖体RNA操纵子之间的关系。【方法】根据亚洲种23S-5S rRNA基因区域序列的保守性设计引物，在非洲种和美洲种DNA样品上扩增，对PCR产物进行克隆和测序，并对新获得的序列进行序列验证和分析。【结果】非洲种获得了3 057 bp 序列包括23S rRNA 基因、细胞壁脱氢酶假基因和5S rRNA 基因；美洲种获得了3 033 bp序列包括23S rRNA 基因、glpK基因和5S rRNA 基因。3个种核糖体基因顺序分别是：亚洲种16S rRNA、tRNAIle、tRNAAla、23S rRNA、细胞壁脱氢酶假基因、5S rRNA 和 tRNAMet；非洲种16S rRNA、tRNAIle、tRNAAla、23S rRNA、细胞壁脱氢酶假基因和 5S rRNA；美洲种16S rRNA、tRNAIle、tRNAAla、23S rRNA、glpK 基因和5S rRNA。【结论】黄龙病病原菌核糖体RNA基因有特殊的排列方式，即在23S rRNA和5S rRNA基因区间均有反向编码的细胞壁脱氢酶基因，其中亚洲种和非洲种为假基因，美洲种为glpK基因。

关键词: 柑橘, 黄龙病, 非洲种, 美洲种, 23S-5S rDNA

Abstract: 【Objective】 The objective of this study is to clone 23S-5S rDNA sequences of Candidatus Liberibacter africanus and americanus and compare to the corresponding region sequences of Candidatus Liberibacter asiaticus to clarify the ribosomal RNA operons relationships of HLB three species.【Method】The HLB DNA samples from africanus and americanus were amplified by the primers designed according to the 23S-5S rRNA gene conserved sequences of asiaticus. The PCR products were cloned and sequenced. The newly obtained sequences were further identified and analyzed.【Result】Sequences of 3 057 bp in length were obtained from africanus including 23S rRNA gene, cell wall hydrolase pseudogene and 5S rRNA gene. Sequences of 3 033 bp in length were obtained from americanus including 23S rRNA gene, glpK gene and 5S rRNA gene. The gene order is 16S rRNA, tRNAIle, tRNAAla, 23S rRNA, cell wall hydrolase pseudogene, 5S rRNA and tRNAMet for asiaticus. 16S rRNA, tRNAIle, tRNAAla, 23S rRNA, cell wall hydrolase pseudogene, 5S rRNA for africanus. 16S rRNA, tRNAIle, tRNAAla, 23S rRNA, glpK gene, 5s rRNA for americanus. 【Conclusion】The operon arrangement was special for three HLB species. The cell wall hydrolase gene coding in the reverse strand was found between 23S rRNA and 5S rRNA gene in three HLB species. It is a pseudogene for asiaticus and africanus while it is glpK gene for americanus.

Key words: Citrus, HLB, Candidatus Liberibacter africauns, Candidatus Liberibacter americanus, 23S-5S rDNA

廖惠红, 李杨瑞, 杨丽涛, 徐宁. 柑橘黄龙病病原菌非洲种和美洲种23S-5S rDNA序列的克隆及分析[J]. 中国农业科学, 2011, 44(17): 3683-3693.

LIAO Hui-Hong, LI Yang-Rui, YANG Li-Tao, XU Ning. 23S-5S rDNA Sequence Cloning and Analysis of Candidatus Liberibacter africanus and americanus of Citrus Huanglongbing[J]. Scientia Agricultura Sinica, 2011, 44(17): 3683-3693.

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柑橘黄龙病病原菌非洲种和美洲种23S-5S rDNA序列的克隆及分析

23S-5S rDNA Sequence Cloning and Analysis of Candidatus Liberibacter africanus and americanus of Citrus Huanglongbing

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