苹果炭疽菌的分子鉴定与检测

测定苹果炭疽菌rDNA全序列，比对苹果炭疽菌和其它炭疽菌ITS序列以及构建系统关系树，发现苹果炭疽菌与胶孢炭疽菌的ITS序列相似性高达99．8％，并与胶孢炭疽菌聚在一起，可以明确苹果炭疽菌应属于胶孢炭疽菌。进一步的序列比对发现，苹果炭疽菌的18S rDNA3’端比其它胶孢炭疽菌多出一段379bp的序列，根据这一特有片段设计引物CgF1与通用引物ITS4配对，结果仅能从苹果炭疽菌中扩增出1232bp的特异性条带。用苹果炭疽菌接种离体苹果，以接种发病的病组织总DNA为模板，利用引物CgF1／ITS4进行PCR扩增，同样可以扩增出1232bp的特异性条带，而健康苹果组织DNA中未能扩增出任何条带，表明该方法可用于苹果炭疽菌的鉴定和快速检测。

Molecular Identification and Detection of Apple Anthracnose Pathogen

The rDNA sequence of apple anthracnose pathogen was determined. Sequence comparing of ITS of CoUetotrichum was conducted and a phylogenetic tree based on alignment of their ITS nucleotide sequences was constructed. The results showed that Cg-1 and Cg-2 shared the highest sequence similarity （99.8 % ） with C. gloeosporioides and they could be grouped into one branch with C. gloeosporioides. It was confirmed that apple anthracnose pathogen should belong to C. gloeosporioides. It was found that there is a 379 bp sequence residue in 3＇ ends of 18S rDNA of apple anthracnose pathogen comparing with the other C. gloeosporioides. Specific primer CgF1 was designed according to the 379 bp sequence, and only a 1 232 bp specific band could be amplified using primer pair CgF1 and ITS4. Apple fruits were inoculated with apple anthracnose pathogen. Total DNA was extracted from the diseased tissue, and an identical 1 232 bp band was amplified. It was suggested that the method could be applied for the detection and rapid identification of apple anthracnose pathogen.