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Method of diagnosing invasive aspergillosis and oligonucleotides used in this method

机译:诊断侵袭性曲霉病的方法和该方法中使用的寡核苷酸

摘要

In the present invention, there is disclosed a method for in vitro diagnostics of invasive aspergillosis by means of isolation and detection of Aspergillus fumigatus DNA in biological samples taken from the body of a patient, said method being characterized in that it contains the following steps: a) isolation of DNA from the clinical samples, b) preamplification of fungal DNA using primers targeting conservative parts of Aspergillus fumigatus ribosomal DNA (rDNA) genes, c) quantitative real-time PCR detection of species specific region of the internal transcribed spacer 2 (ITS2) part of rDNA genes of an Aspergillus species wherein the step c) is performed using as forward primer an oligonucleotide having at least 85 percent homology to the sequence GGCTTTGTCACCTGCTCTGTAG (SEQ ID NO.2), as reverse primer an oligonucleotide having at least 85 percent homology to the sequence CTGATCCGAGGTCAACCTTAGAAA (SEQ ID NO.3) and as a probe an oligonucleotide having at least 85% homology to a sequence selected from the group comprising CCGACACCCAACTTT û MGB (SEQ ID NO.4) wherein the reduction in homology down up to 85 percent is achieved by extension or shortening of the primers and/or the probe. There are also disclosed novel oligonucleotides intended for the use in invasive aspergillosis diagnostics.
机译:在本发明中,公开了一种通过分离和检测从患者体内获取的生物样品中的烟曲霉DNA来体外诊断侵袭性曲霉病的方法,所述方法的特征在于它包含以下步骤: a)从临床样本中分离DNA,b)使用靶向烟曲霉核糖体DNA(rDNA)基因保守部分的引物对真菌DNA进行预扩增,c)实时定量PCR检测内部转录间隔区2的物种特异性区域曲霉属物种的rDNA基因的ITS2)部分,其中步骤c)是使用与序列GGCTTTGTCACCTGCTCTGTAG(SEQ ID NO.2)具有至少85%同源性的寡核苷酸作为正向引物进行的,作为反向引物是具有至少85与序列CTGATCCGAGGTCAACCTTAGAAA(SEQ ID NO.3)的百分比同源性,作为探针的与序列至少具有85%同源性的寡核苷酸选自CCGACACCCAACTTT MGB(SEQ ID NO.4)的组,其中同源性降低至多85%是通过延伸或缩短引物和/或探针来实现的。还公开了旨在用于侵入性曲霉病诊断的新型寡核苷酸。

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