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【目的】柳树黄化病是一种重要的植原体病害,本研究旨在明确柳树黄化病植原体(Willow Yellow phytoplasma,WY)的分类地位,为进一步开展致病性和防治研究奠定基础。【方法】采用植原体特异引物通过PCR方法从患病植株DNA中扩增植原体16S rDNA基因和核糖体蛋白基因(ribosomal proteins gene,rp),对所得的序列进行分析,构建同源进化树,并用限制性片段长度多态性(RFLP)对巢式PCR产物进行分析。【结果】首次从柳树黄化病植原体中分离出了16S rDNA基因和rp基因,大小分别为1246bp和1212bp。通过对植原体16S rDNA和rp基因的核苷酸同源性比较和RFLP分析,发现该分离物与16SrI组的核苷酸同源性均在99%以上,与16SrI-C亚组中的小麦蓝矮病植原体同源性高达99.8%(16SrDNA)和99.6%(rp),且RFLP分析与16SrI-C亚组的植原体有相同的酶切条带。【结论】柳树黄化植原体应划分于16SrI-C亚组。
【Objective】 Willow yellow disease is one of the important phytoplasma diseases. The purpose of this study is to clarify the taxonomic status of willow yellow phytoplasma (WY) and lay a foundation for further study on pathogenicity and control. 【Method】 Phytoplasma 16S rDNA gene and ribosomal proteins gene (rp) were amplified from diseased plants by PCR using phytoplasma-specific primers. The sequences were analyzed to construct homologous phylogenetic tree, The nested PCR products were analyzed by restriction fragment length polymorphism (RFLP). 【Result】 The results showed that 16S rDNA gene and rp gene were isolated from the willow yellow phytoplasma for the first time with sizes of 1246bp and 1212bp, respectively. By comparing the nucleotide homologies of 16S rDNA and rp genes with the RFLP analysis of phytoplasmas, the nucleotide homologies of the 16S rI and 16SrI isolates were all above 99% Blue dwarf phytoplasmas have 99.8% (16S rDNA) homology and 99.6% (rp) homology with the RFLP analysis. The RFLP analysis showed the same restriction bands as the 16SrI-C subgroup. 【Conclusion】 Salix williatus should be divided into 16SrI-C subgroups.