新疆枣疯病植原体tuf基因的克隆与序列分析

Cloning and Sequencing Analysis of tufGene of the P hytoplasma Associated with Jujube Witches Broom in Xinjiang

【目的】枣疯病是一种重要的植原体病害,了解新疆枣疯病植原体的系统发育关系及遗传分化,获取其分子特征信息。【方法】利用植原体tuf基因通用引物fTufu/rTufu对新疆枣疯病病株总DNA进行PCR扩增,对部分扩增片段克隆、测序及相似性分析。【结果】获得新疆枣疯病阿克苏株系(JWB-ASZ)和喀什株系(JWB-KHZ)的tuf基因片段分别为841和814 bp。经同源性比较分析,新疆枣疯病阿克苏株系(JWB-ASZ)与喀什株系(JWB-KHZ)均隶属于16S rⅤ-B亚组枣疯病植原体。其中JWB-KHZ株系与枣疯病植原体河北株系的tuf基因核苷酸及氨基酸同源性最高,分别达到90.2%和81.5%;JWB-ASZ与枣疯病植原体陕西株系的tuf基因核苷酸及氨基酸同源性最高,分别达到94.2%和94.6%,而与16S rⅤ其它亚组的核苷酸及氨基酸同源性均低于70%。枣疯病阿克苏株系与喀什株系的tuf基因核苷酸及氨基酸序列之间存在明显差异,同源性仅为为73.5%和63.5%。【结论】研究报道了新疆枣疯病植原体tuf基因序列,为揭示新疆枣疯病植原体不同株系的分子特征、分子变异和遗传多样性提供了基础。

[ Objective ] Jujube witches broom （JWB） is an important plant disease caused by phytoplasma. The major objective of our research was to realize the molecular phylogenetie relationship, genetic differentiation and obtain molecular characteristics of JWB phytoplasmas of Xinjiang isolates. [ Method ] By using of PCR, the elongation factor Tu （ tuf gene） of phytoplasma associated with jujube witches broom in the total DNA extracted from diseased samples in Aksu and Kashi region of Xinjiang was amplified with the universal primer pairs frufu/rTufu, then the partial tuf gene was cloned and sequenced, and similarity to other phytoplasmas was analyzed. [ Result ] The sequencing result revealed that tuf gene of phytoplasmas from JWB - ASZ strain isolated in Aksu was 841 bp in length and JWB - KHZ strain isolated in Kashi was 814 bp in length. Homology analysis of sequences of tuf gene showed that the JWB - ASZ and JWB - KHZ were closely related to those in the 16S rV -B subgroup of jujube witches＇ broom phytoplasma. In tuf gene, JWB -KHZ strain isolated in Kashi shared most similarity with JWB strain Hebei which had been already reported, and homology rates of nucleotide sequences and amino acids were 90.2% and 81.5%, respectively. JWB - ASZ strain isolated in Aksu shared most similarity with JWB in Shaanxi district, and homology rates of nucleotide sequences and amino acids were 94.2% and 94.6%, respectively. However, they showed a low similarity （ 〈70% } with members of the other 16S rV subgroup phytoplasmas. And the remarkable differences of the tuf gene nucleotide and the derived amino acid sequence similarities between the two isolates were 73.5% and 63.5%. [ Conclusion ] This paper for the first time reports the tuf gene sequences of J-WB phytoplasmas of Xinjiang isolates, thus directly providing a theoretical basis for research of the phytoplasma molecular characterization, molecular variation and genetic diversity of the different phytoplasmas associated with JWB in Xinjiang.