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本研究选取2011年采自河南烟区的32个菌株,根据4个核基因(包括rDNA ITS、β-tubulin、Ras和Gpi基因)和1个线粒体基因(Cox1)的保守序列,克隆和测序分析160个相应的基因DNA片段。结果显示,这5个基因DNA序列在病菌群体内碱基序列与参照基因序列相比保守度在83%~100%不等,平均基因型多样性指数(M)为1.422 7。综合使用5个基因联合序列联合构建群体进化系统树,揭示了分布于河南省14个县区的32个分离菌系可分为2个明显的组,包括15个不同的联合基因型,基因型多样性指数为2.35,反映了病菌致病变种内存在的遗传变异多样性。其中,最优势的2个联合基因型分别涵盖8个c基因型菌系和6个e基因型菌系。然而,各基因型间遗传分化的总水平很低,组间的遗传距离仅为0.000 5,说明烟草黑胫病菌在河南生长烟草中的菌株可能属于同一有效群体。同时,不同分化菌株遗传亲缘关系与地理来源之间也没有显著的相关性,表明烟草黑胫病菌河南群体内存在密切的菌源交流,没有形成明显的地理亚群体分化。本研究结果可广泛应用于大规模黑胫病菌群体菌系的动态检测,服务于烟草抗黑胫病育种和品种合理布局。
Based on the conserved sequences of four nuclear genes (including rDNA ITS, β-tubulin, Ras and Gpi genes) and one mitochondrial gene (Cox1), 32 clones were selected from Henan tobacco growing areas in 2011, cloned and sequenced 160 corresponding gene DNA fragments. The results showed that the DNA sequences of these 5 genes had a conservative degree of 83% -100% compared with the reference gene sequences in the bacterial population. The average genotype diversity index (M) was 1.4227. Using a combination of five gene sequences to construct a phylogenetic tree, it was revealed that 32 isolates distributed in 14 counties of Henan province could be divided into two distinct groups, including 15 different combinations of genotypes, genotypes The diversity index was 2.35, reflecting the diversity of genetic variation found in pathogenic strains of pathogens. Among them, the two most favorable combinations of genotypes include 8 c genotypes and 6 e genotypes, respectively. However, the overall level of genetic differentiation among the genotypes was very low, and the genetic distance between the two groups was only 0.0005, indicating that the strains of Phytophthora capsici in tobacco growing in Henan may belong to the same effective group. At the same time, there was no significant correlation between genetic relationship and geographic origin of different strains, indicating that there is close bacterial source exchange in Henan population of Phytophthora nicotiana and no obvious differentiation of geographical subgroups. The results of this study can be widely used in the dynamic detection of large-scale blackleg genotype bacterial strains, serving the anti-blackleg disease breeding and variety reasonable layout.