中国各地不同枣树品种上枣疯病植原体的PCR检测及分子变异分析

摘要：【目的】检测不同地区枣树品种上的枣疯植原体侵染及保守基因序列的变异。【方法】利用植原体16S rDNA的通用引物R16mF2／R16mR1、16S-23S间区序列(SR)的通用引物SR1/SR及secY基因引物FD9f/r，通过PCR检测采自国内7个地区14个枣树品种上的32个枣疯病和4个酸枣丛枝病样品。将PCR产物进行直接或克隆测序，结合已报导的测序数据，进行序列同源性和系统进化分析。【结果】所有枣疯病样品中均检测到植原体；皆属于榆树黄化16S rV-B亚组，与我国重阳木丛枝和樱桃致死黄化遗传关系

Molecular detection and variability of jujube witches'-broom phytoplasmas from different cultivars in various regions of China

Objective] Jujube witches'-broom is an important disease in jujube cultivation areas, which causes serious losses in jujube fruit production. To understand the genetic variability and diversity of jujube witches'-broom phytoplasma population from the different cultivars and various regions of China. Method] We collected 32 samples from 14 cultivars or wild sour jujubes in 7 regions of China and detected them with PCR with the primers RI6mF2/R16mR1 for phytoplasma 16S rDNA, SRI/SR for16S23SrRNA space region (SR) and FDgf/r for secretion proteins (secY). The direct sequencing of PCR products and sequencing by cloned PCR products were used for sequence polymorphism and phylogenetic analyses by comparison to the databases of known conserved gene sequences. Results] We detected phytoplasmas by PCR amplification of 16SrDNA from all the diseased jujube samples. All the phytoplasma isolates infected various jujube cuhivars belonged to subgroup 16SrV-B of elm yellows group and had closer homology with Bischofia polycarpa witches'-broom and cherry lethal yellows phytoplasmas occurred in China than other 16SrV phytoplasmas in other countries. The sequence polymorphism at different extent in 16SrDNA, SR and secY gene and genetic diversity were revealed in phytoplnsma strain population related to different habitats, among which the dominant strains were always detected by the direct sequencing of PCR products in all the diseased areas of China. The degree of variability on secY gene of collected phytoplasma strains was greater than that of 16SrDNA and SR sequences, and some base substitutions could not alter encoded amino acid, however certain single base deletions detected in a Shandong and a Beijing strains may have impact on the gene structure or function. Conclusion] Phytoplasma strains from different cultivars and regions show dramatic genetic diversity. Compared with direct sequencing of PCR products, the sequencing by cloning PCR products was more useful for the displaying of variants and phylogeny in phytoplasma strain population.