海南竹柏扁枝病病原植原体的分子检测鉴定

Molecular Identification of a Phytoplasma Associated with Podocarpus nagi Fasciation in Hainan

提取采自海南万宁热带植物园中表现典型扁枝症状的竹柏植株总DNA,利用植原体16S r RNA基因通用引物对P1/P7和R16F2n/R16R2进行巢式PCR检测。结果表明:扩增到大小约1.2 kb的植原体特异片段,通过对扩增片段进行克隆、测序和序列比对分析,结果确定为植原体感染。系统进化分析结果表明,该植原体(Gen Bank登录号:KP027298)为australasiae植原体候选种相关株系,与australasiae植原体候选种(Gen Bank登录号:Y10097)的同源性为99.4%。进一步虚拟RFLP分析,结果表明该植原体属于花生丛枝植原体组(16Sr II)的一个新亚组,与其相似性最高的是16Sr II-A亚组(相似系数为0.97)。为指导竹柏扁枝病的防治奠定理论依据。

In this paper, universal primers of P1/P7 and R16F2n/R16R2 for phytoplasmal 16S rRNA gene were used to detect phytoplasma by nested PCR respectively. The 1.2 kb DNA fragments were amplified from the total DNA of Podocarpus nagi （Thunb.） Zoll. et Mor ex Zoll with fasciation disease symptoms, which were collected from the Tropical Botanical Garden, Wanning, Hainan. Sequence analysis determined that the P. nag/ with fasciation disease was caused by a phytoplasma（GenBank accession： KP027298）, which shared 99.4% similarity of the ＇ Candidatus Phytoplasma australasiae＇reference strain （GenBank accession： Y10097）. Furthermore, virtual RFLP analysis showed that the phytoplasma belonged to to a new subgroup within the peanut witches＇-broom （16Sr II） group, the most similar was the reference pattern of the 16Sr group II, subgroup A（GenBank accession： L33765）, with a similarity coefficient of 0.97. The study is important for effective prevention of the disease.