Universal fungus-specific primer systems and group-specific hybridization oligonucleotides for 18S rDNA
Universal fungus-specific primer systems and group-specific hybridization oligonucleotides for 18S rDNA
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DOI：
10.1111/j.1439-0507.1996.tb00079.x
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发表时间：
1996-01-01
期刊：
影响因子：
4.9
通讯作者：
Maiwald, M
中科院分区：
文献类型：
--
作者：
Kappe, R;Fauser, C;Maiwald, M
我们设计了两个引物系统，扩增编码核糖体小亚基（18S rRNA）的基因片段。一种广泛反应性但真菌特异性的引物混合物包括两种先前公布的特异性扩增皮肤真菌的引物TR1和TR2，以及两种新设计的分别特异性扩增念珠菌和曲霉的引物CA1和AF2。该引物混合物扩增长度约为578个碱基对（bp）的DNA片段（从位置838至1415），其含有可变的、可能是物种特异性的区域（V5，部分V7）。另一种新设计的引物UF 1（通用真菌引物1）沿着真核引物S3扩增了一个926 bp的片段（从263位到1188位），该片段包括可变区V3、V4和V5。两种引物系统都扩增了酿酒酵母、白色念珠菌、新型隐球菌、烟曲霉、马尔尼菲青霉、尖孢镰刀菌和须癣毛癣菌的DNA，但不能扩增佐氏无绿藻、大肠杆菌或人的DNA。用已发表的对皮肤癣菌和组织胞浆菌特异的寡核苷酸TR和HC以及新设计的组特异寡核苷酸CA和AF与T.须癣菌、荚膜组织胞浆菌、C. albicans和A.对其他6种真菌和3种对照均无显著影响。
We designed two primer systems that amplify a fragment of the gene coding for the small ribosomal subunit (18S rRNA). A broadly reactive, yet fungus-specific, primer cocktail comprises two previously published primers, TR1 and TR2, which specifically amplify dermatophytes, and two newly designed primers, CA1 and AF2, which specifically amplify Candida and Aspergillus respectively. This primer cocktail amplifies a DNA fragment of approximately 578 basepairs (bp) in length (from position 838 to 1415), which contains variable, possibly species-specific regions (V5, partly V7). Another newly designed primer, UF1 (universal fungal primer 1), along with the eukaryotic primer S3 amplifies a 926-bp fragment (from position 263 to 1188) that includes the variable regions V3, V4 and V5. Both primer systems amplified DNA from Saccharomyces cerevisiae, Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, Penicillium marneffei, Fusarium oxysporum and Trichophyton mentagrophytes, but not the DNA from Prototheca zopfii, Escherichia coli or humans. The previously published oligonucleotides TR and HC, which are specific for dermatophytes and Histoplasma respectively, and the newly designed group-specific oligonucleotides, CA and AF, hybridized with T. mentagrophytes, Histoplasma capsulatum, C. albicans and, A. fumigatus respectively, but not with the other six fungi or with the three controls.