橡胶树尖孢炭疽菌分子检测及遗传多态性分析

摘要

根据GeneBank中Colletotrichum属的20个种的ITS序列,比较设计出1对引物CAF53/CAR356(CAF53 5'-GGG CAG GGG AAG CCT CTC G-3';CAR356 5'-ACC GGT GCT TGA GGG TTG-3');该引物可以扩增出1条303 bp的尖孢炭疽菌特异性DNA条带.并利用ISSR和RAPD分子标记技术对从海南、广东、广西和云南等垦区橡胶树上收集的21个尖孢炭疽菌株进行遗传多态性分析,以杧果上分离的尖孢作为参照菌株进行了聚类分析.结果表明,RAPD的平均遗传相似系数为0.8,ISSR的平均遗传相似系数为0.7,2种分子标记的平均遗传相似系数基本一致,均可揭示尖孢炭疽病菌的遗传多态性;2种分子标记产生的聚类分析结果存在一定差异:ISSR和RAPD类群与菌株的地理来源均不存在相关性.%Based on the differences of ITS DNA sequences of 20 species of Collewtrichum from GenBank, a couple of specific primers of CAF53/CAR356(CAF53: 5'-GGG CAG GGG AAG CCT CTC G-3'; CAR356: 5'-AGC GGT GCT TGA GGG TTG-3') were designed to detect Colletotrichum acutatum by PCR. A 303 bp DNA fragment could be amplified only from C. acutatum genomic DNA with CAF53/CAR356 primers under the optimized PCR parameters.The picogram level of genomic DNA of C. acutatum could be detected by this approacl. 22 isolates of Collewtrichum acutatums were analyzed with random amplified polymorphic DNA(RAPD) and internal simple sequence repeat (ISSR) methods. The results showed that the two molecular markers had consistent genetic coefficient of similarity and beth could revealed the genetic diversity of isolates in C. acutatum. The cluster results produced by the two techniques were different. The RAPD and ISSR groups were not associated with geographic regions of isolates.