title: [Development and application of TaqMan MGB probe fluorescence quantitative PCR method for rapid detection of Clostridium piliforme].
abstract: OBJECTIVE
  To develop a TaqMan MGB probe-based, sensitive and specific fluorescence quantitative PCR assay method for rapid detection of Clostridium piliforme.
  
  
  METHODS
  Primers and probes specific to 16S rRNA gene of Clostridium piliforme were designed. A TaqMan MGB probe-based, fluorescence quantitative PCR method was established. Specificity, sensitivity and stability of the method were assessed, followed by real-time quantitative PCR assay to detect Clostridium piliforme on 1156 clinical specimens during 2008-2011 and compared with conventional PCR assay.
  
  
  RESULTS
  The specificity of TaqMan MGB probe-based fluorescence quantitative PCR was high and did not show cross-reactivity with Helicobacter hepaticus, Helicobacter pylori, Campylobacter jejuni, Pasteurella pneumotropica, Escherichia coli or Pseudomonas aeruginosa. The detection limit was 2.2 copies/µl. The correlation coefficient and slope value of standard curve were 0.999 and -3.204, respectively and the efficiency of TaqMan MGB-based probe fluorescence quantitative PCR assay was 100%. When the TaqMan MGB-based probe fluorescence quantitative PCR assay was preformed to detect Clostridium piliforme on 1156 clinical specimens, a total of 101 specimens showed positive on Clostridium piliforme. However, only 44 specimens showed positive when conventional PCR was used. The real-time quantitative PCR for Clostridium piliforme could be completed within 2 hours.
  
  
  CONCLUSIONS
  The TaqMan MGB-based probe fluorescence quantitative PCR assay method was a reliable, specific, sensitive and useful tool for rapid detection of Clostridium piliforme.
authors.affiliation: National Institutes for Food and Drug Control of China, Beijing, China. gaozhengqin@126.com
authors.forename: Zheng-qin
authors.initials: ZQ
authors.lastname: Gao
authors.forename: Bing-fei
authors.initials: BF
authors.lastname: Yue
authors.forename: Zheng-ming
authors.initials: ZM
authors.lastname: He
mesh_terms: D003013:Clostridium / Q000302:isolation & purification*; D006801:Humans; D015341:Nucleic Acid Probes; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D015341:Nucleic Acid Probes