摘要
目的:利用形态学特征与Nested-PCR技术鉴定黄檗丛枝菌根真菌。方法:采用酸性品红染色法挑选黄檗丛枝菌根。同时,利用湿筛法获得AM真菌孢子,进行形态学鉴定。运用Nested-PCR技术,对黄檗粗提DNA进行特异性扩增,采用blastn进行序列相似性比较,并构建系统进化树,确定侵染黄檗根系的AM真菌。结果:编号为HDAM-1的AM真菌孢子,形态特征与G.intraradices的特征描述一致。Nested-PCR检测到约455bp的目的片段,其序列与G.intraradices(DQ469118)相似性最高,达97.8%,有11个碱基的差异。系统进化树显示该序列在基于25SrDNA的进化树中与G.intraradices(DQ469118.1)处于同一分支,确定G.intraradices侵染黄檗根系。结论:将形态学特征与Nested-PCR技术相结合鉴定AM真菌,不仅简易、经济,而且能够提高研究结果的可靠性。
Objective： To identify the arbuscular mycorrhizal fungi of Phellodendren amurense Rupr. by Morphological Characteristics and Nested - PCR. Methods： The arbuscular mycorrhizal of Phellodendren amurense Rupr. were detected by acid fuchsin staining. At the same time, many AM fungal spores were accessed by Wet- screening method and identified by their Morphological Characteristics. The crude DNA extracted from Phellodendren amurense Rupr. were applied to Nested- PCR by specific primers. To detemaine the AM fungi of Phellodendren amurense Rupr., the sequence of the target fragment was compared with the known sequences in the GenBank database. Then, Phylogenetic tree was constructed by analysis of sequences of the known species. Results： The spores of an arbuscular mycorrhizal fungi marked as HDAM- 1 were in accord with G. intraradices in morphology. The target fragment（455bp）was detected by Nested- PCR. The target fragment was closely related to intraradices（DQ469118）, with the similarity of 97.8%. There is only 11 bases in 25S rDNA region of HDAM - 1 different from Glomus intraradices （ DQ469118 ）. The phylogenetic analysis showed that HDAM - 1 come together in Glomus intraradices （DQ469118 ） on the basis of 25S rDNA sequence. It showed that Phellodendren amurense Rupr. was infected by G. intraradices. Conclusion： Identification of the AM fungi by Morphological Characteristics and Nested- PCR is simple, economical, and the reliability of findings was improved.
出处
《生物技术》
CAS
CSCD
2007年第6期32-35,共4页
Biotechnology
基金
黑龙江省博士后基金项目(LBH-Z05013)
黑龙江大学博士启动基金项目资助
关键词
黄檗
丛枝菌根真菌
酸性品红染色
湿筛法
鉴定
Nested—PCR
Phellodendren amurense Rupr.
arbuscular mycorrhtizal fungi
acM fuchsin staining
wet - screening method
identification
Nested -PCR
