title: Differentiation of fungi using hybridization probes on the LightCycler(®).
abstract: The polymerase chain reaction is a powerful molecular tool for the detection and analysis of very small amounts of DNA. Today, hybridization probes are often used in real-time PCR for more sensitive and specific detection of pathogens and for determination of gene regulation or mutation analysis instead of intercalating dyes like SYBR Green. Here, we describe how to generate suitable primers and hybridization probes for the specific detection of fungal DNA. Furthermore, we show the advantages of hybridization probes using the LightCycler-PCR for the detection of different Candida spp. and Aspergillus spp. in patient blood samples. The methods used to develop such PCR assays will also be presented in the following protocol.
authors.affiliation: Fraunhofer Institute for Cell Therapy and Immunology (IZI), Leipzig, Germany. stephan.fricke@izi.fraunhofer.de
authors.forename: Stephan
authors.initials: S
authors.lastname: Fricke
authors.forename: Nadja
authors.initials: N
authors.lastname: Hilger
authors.forename: Christopher
authors.initials: C
authors.lastname: Oelkrug
authors.forename: Arne C
authors.initials: AC
authors.lastname: Rodloff
authors.forename: Christian
authors.initials: C
authors.lastname: Fricke
mesh_terms: D001230:Aspergillus / Q000235:genetics* / Q000302:isolation & purification*; D002175:Candida / Q000235:genetics* / Q000302:isolation & purification*; D017931:DNA Primers / Q000235:genetics; D005456:Fluorescent Dyes / Q000737:chemistry*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D015345:Oligonucleotide Probes / Q000737:chemistry / Q000235:genetics*; D012331:RNA, Fungal / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D013045:Species Specificity
chemical_list: D017931:DNA Primers; D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes; D012331:RNA, Fungal; D012337:RNA, Ribosomal, 18S