Comparison of TaqMan™ and Epoch Dark Quenchers™ during real-time reverse transcription PCR

• Published in: Molecular and cellular probes 2004-06, Vol.18 (3), p.207-209
• Main Authors: Daum, Luke T, Ye, Keying, Chambers, James P, Santiago, Jose, Hickman, John R, Barnes, William J, Kruzelock, Russell P, Atchley, Daniel H
• Format: Article
• Language: English
• Subjects: Dark Quenchers; Fluorescent Dyes - metabolism; Influenza A virus - genetics; Real-time fluorogenic PCR; Reverse Transcriptase Polymerase Chain Reaction - instrumentation; Reverse Transcriptase Polymerase Chain Reaction - methods; RNA, Viral - genetics; Sensitivity and Specificity; Taq Polymerase - metabolism; TaqMan PCR
• ISSN: 0890-8508; 1096-1194
• DOI: 10.1016/j.mcp.2004.01.005

Several biotechnology companies have recently introduced novel quencher fluors for use with dual-labeled fluorogenic hydrolysis probes. The Epoch Dark Quencher™ fluorochrome consists of a non-fluorescent moiety capable of absorption at higher wavelengths (400–650 nm). The aim of this study was to: (1) evaluate the feasibility of using Epoch Dark Quencher™ fluorochromes in real-time PCR pathogen detection assays that were previously optimized with TaqMan™ (TAMRA) quenching fluors, and (2) compare the sensitivity based on cycle threshold ( C T) between probes containing either TaqMan™ or Epoch Dark Quencher™ fluors. Our data indicate Epoch Dark Quencher™ probes can be used in place of TaqMan™ probes and their performance was not better than traditional TaqMan (TAMRA) quenchers. Marginal differences observed between quenching fluorochromes may arise from concentration differences during probe synthesis.