刘金华,贺丹,史艳宇,黄宇,王丽,任常菲
LIU Jin-hua△, HE Dan, SHI Yan-yu, et al (△Department of Pathogenic Biology, College of Basic Medicine, Jilin University

• 1:吉林大学基础医学院病原生物学教研室
• 2:吉林出入境检验检疫局
• 3:吉林省产品质量监督检验院

摘要(Abstract)：

目的建立黑曲霉实时荧光PCR检测方法。方法对6种主要病原曲霉(黑曲霉、烟曲霉、杂色曲霉、构巢曲霉、土曲霉及黄曲霉)的GAPDH基因序列进行比对分析,选择黑曲霉特异位点设计引物和探针,对黑曲霉进行实时荧光PCR扩增,并检测该方法的灵敏度及特异性。结果该方法可检出2.78×10-10μg/ml的黑曲霉基因组DNA;对亲源关系较近的11株不同种曲霉及4株其他属临床常见的病原真菌进行实时荧光PCR检测,未发现有交叉反应。结论已建立了灵敏度高、特异性好的快速检测黑曲霉的实时荧光PCR方法。
Objective To develop a real-time fluorescent PCR for determination of Aspergillus niger. Methods The GAPDH gene sequences of 6 kinds of pathogenic Aspergillus, i.e. Aspergillus niger, Aspergillus fumigatus, Aspergillus versicolor, As-pergillus nidulans, Aspergillus terreus and Aspergillus flavus, were compared, based on which primers and probes for real-time fluo-rescent PCR were designed according to the specific site of Aspergillus niger. The developed real-time fluorescent PCR was analyzed for sensitivity and specificity. Results By using the developed real-time fluorescent PCR, 2. 78 × 10-10 μg / ml of genomic DNA of Aspergillus niger was detected. Eleven Aspergillus niger isolates with close genetic relationship and 4 isolates of other common pathogenic fungi in clinic were determined by the developed method, and no cross reactions were observed. Conclusion A real-time fluorescent PCR for rapid determination of Aspergillus niger, with high specificity and sensitivity, was developed.

关键词(KeyWords)： 黑曲霉;实时荧光PCR;特异性;灵敏度
Aspergillus niger; Real-time fluorescent PCR; Specificity; Sensitivity

Abstract:

Keywords:

基金项目(Foundation): 吉林省科技发展计划重点资助项目(20080444-2)

作者(Author): 刘金华,贺丹,史艳宇,黄宇,王丽,任常菲
LIU Jin-hua△, HE Dan, SHI Yan-yu, et al (△Department of Pathogenic Biology, College of Basic Medicine, Jilin University

DOI: 10.13200/j.cjb.2009.03.88.liujh.024

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