Real Time/Quantitative PCR Competence and Service in Molecular Biology

Real Time/Quantitative PCR metabion offers Black Hole series of dark quenchers (BHQTM) which were designed for use in real-time PCR assays. These molecules function as efficient quenchers of fluorescence over the entire visible spectrum and into the IR, re-emitting their energy as heat rather than light. Probes made with BHQ quenchers exhibit extremely low background noise and allow for lower copy sensitivity. BHQ-1 has its max absorption at 534 nm and quenches most efficently in the 480 to 580 nm range. BHQ-2 has its λmax at 579 nm, which enables it to “absorb” fluorescence in the 550 to 650 range. BHQ-3 has its max absorption at 672 nm with an optimal quenching range of 620-730 nm. Competence and Service in Molecular Biology

Real Time/Quantitative PCR BHQ Dye Absorption Maxima and Quenching Range Quencher Abs (nm) Quenching Range (nm) BHQ-1 534 480-580 BHQ-2 579 550-650 BHQ-3 672 620-730 BHQ Dye / Reporter Combinations Quencher Suggested Fluorophores BHQ-1 FAM, TET, JOE, HEX, … BHQ-2 TAMRA, ROX, CY3, LCRed 640, … BHQ-3 CY5.5, IRDye700, LCRed 705, … The unique characteristics of the BHQ classof quenchers permits flexibility in the choice of spectrally well-resolved fluorophores enabling single-tube multiplexing with little or no cross-talk Competence and Service in Molecular Biology

Real Time/Quantitative PCR Absorption spectra of DABCYL-T9 and BHQ-1-T9 with the emission spectra of FAM, TET, and JOE. DABCYL and BHQ-1 dye were normalized at the poly-T-absorbance (i.e. 260 nm) to better demonstrate the larger extinction coefficient of the BHQ-1 dye. RFU=relative fluorescence units. Competence and Service in Molecular Biology