荧光定量PCR检测霉变烟叶中米曲霉方法的建立  

Establishment of a Real-time PCR Assay for Detection of Mold Fungi in Tobacco

作　　者：蓝东明[1] 唐庆芸[1] 杨博[2] 王永华[1] 

机构地区：[1]华南理工大学轻工与食品学院,广东广州510640 [2]华南理工大学生物科学与工程学院,广东广州510006

出　　处：《现代食品科技》2013年第7期1721-1724,共4页Modern Food Science and Technology

基　　金：广州市科技发展重点项目(粤烟221201II科字第15号)

摘　　要：为建立快速高效的烟草霉变微生物定量与定性的检测方法,本研究针对陈旧烟叶中分离的米曲霉的rDNAITS1基因片设计一对特异引物,制备含ITS1基因的重组质粒作为阳性对照,建立了米曲霉基因组的SYBR Green I应该定量PCR检测方法。结果显示,标准曲线的相关系数为0.998,最低可检测浓度为101copy/μL的阳性质粒,与其他的微生物的基因组不发生交叉反应,该技术可为检测烟叶霉变微生物提供快速可靠的检测手段。To establish a rapid and effective detection method for mold fungi in tobacco,a real-time PCR assay based on SYBR Green I was developed with a pair of primers designed according to the rDNA ITS1 gene of Aspergillus oryzae isolated from commercial tobacco.A recombinant T-vector containing the ITS1 gene was constructed as a positive control.The correlation coefficient of the standard curve was 0.998 and was highly sensitive with a detection limit of 101copy/μL of positive recombinant plasmid.The technology was specific toward the genome of Aspergillus oryzae without any cross-reaction with other microbe genome.The developed real-time PCR method in this study can be used for detection of mold fungi in tobacco.

关 键 词：丝状真菌 荧光定量PCR 烟草 

分 类 号：TS47[农业科学—烟草工业] O657.3[理学—分析化学]