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PCR TECHNIQUES FOR DETECTING MICROBIAL AND VIRAL CONTAMINANTS IN FOODSTUFFS

机译:检测食品中微生物和病毒污染物的PCR技术

摘要

A method or detecting the presence of living or dead microorganisms and viruses in a sample comprises adding to a pre-determined volume of a sample comprising nucleic acid-containing microbe (s) and/or virus (es), known amounts of a pair of primers binding to sequences up-stream and down-stream to a universal or specific microbial and/or viral nucleic acid sequence and polymerase chain reaction (PCR) reagents, cycling the mixture to amplify the universal or specific microbial and/or viral nucleic acid sequence; adding a polynucleotide comprising a DNA internal segment that is hybridizably complementary to at least a portion of the universal or specific nucleic acid sequence; and a first and a second DNA arm segment adjoining the DNA internal segment, the first DNA arm segment ending in a 5' terminus and the second DNA arm segment ending in a 3' terminus, the arms segments comprising nucleotide sequences such that they are hybridizably complementary to one another. Optionally, the terminus of one arm segment is operatively connected to a fluorogen and the terminus of the other arm segment is operatively connected to a quencher, such that the polynucleotide comprises a "fluorescent beacon."
机译:一种检测样品中是否存在活的或死的微生物和病毒的方法,包括将预定量的包含已知量的含核酸的微生物和/或病毒的样品添加到预定体积的样品中,所述样品包括含核酸的微生物和/或病毒。与通用或特定微生物和/或病毒核酸序列上游和下游序列结合的引物和聚合酶链反应(PCR)试剂​​,循环混合物以扩增通用或特定微生物和/或病毒核酸序列;加入包含与通用或特异性核酸序列的至少一部分可杂交互补的DNA内部区段的多核苷酸;以及与DNA内部片段邻接的第一和第二DNA臂片段,第一DNA臂片段终止于5'末端,第二DNA臂片段终止于3'末端,这些臂片段包含核苷酸序列,使得它们可杂交彼此互补。任选地,一个臂段的末端可操作地连接至氟,而另一臂段的末端可操作地连接至淬灭剂,以使多核苷酸包含“荧光信标”。

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