3rd July 2024

PerfeCTa SYBR Green FastMix for iQ

PerfeCTa® SYBR® Green FastMix for iQ™ is a 2X concentrated, ready-to-use reaction cocktail that contains all components, except primers and template for real-time quantitative PCR (qPCR) on Bio-Rad iCycler iQ™ systems. This unique combination of proprietary buffer, stabilizers, and AccuFast™ Taq DNA polymerase deliver maximum PCR efficiency, sensitivity, specificity and robust fluorescent signal using fast, or conventional, cycling protocols with SYBR® Green qPCR. Highly specific amplification is crucial to successful qPCR with SYBR® Green I dye technology because this dye binds to and detects any dsDNA generated during amplification. AccuFast Taq DNA polymerase is a key component of this reaction mix. This hot-start Taq contains a proprietary mixture of monoclonal antibodies that bind to the polymerase and keep it inactive prior to the initial PCR denaturation step (> 48 hours at room temperature). Similar to our AccuStart Taq DNA polymerase, these antibodies are irreversibly inactivated during the initial PCR denaturation step. However, unlike other antibody hot-start polymerases, activation of AccuFast Taq is instantaneous at 95°C. Rapid recovery of fully active, unmodified Taq DNA polymerase is critical for efficient extension kinetics. Replication of fragments up to 200 bp is complete in less than 20s at 60°C.

  • 2X concentrated master mixes with stabilized SYBR Green dye and exceptional temperature stability (≥30 days at 22°C) that withstand repetitive freeze-thaw (≥ 20X).
  • Rigorously optimized for both fast and conventional thermal cycling parameters.
  • Supports efficient vortex mixing with proprietary anti-foaming technology.
  • Maximizes assay sensitivity and target precision with highly modified Taq DNA polymerase and stringentultrapure, AccuFast™ II antibody hotstart technology.
  • Easy-to-use with AccuVue™ plate loading dye and pre-blended passive reference dye.

Real-Time PCR Instrument Compatibility

  • BioRad iCycler iQ™
  • BioRad MyiQ™
  • BioRad iQ™5

Performance Data

Comparison to Finnzymes, ADAR

PageW - Graph ADAR - 5'ADAR
PerfeCTa SYBR Green FastMix Comparison to DyNAmo Flash SYBR Green PCR Kit
RNA-specific adenosine deaminase (ADAR) was amplified from log-fold dilutions of total HeLa cell cDNA (100 ng to 10 pg) using PerfeCTa™ SYBR Green FastMix or the DyNAmo Flash SYBR Green PCR Kit (Finnzymes) according to each manufacturers protocol. Averaged plots for quadruplicate reactions for each input quantity are shown. Fusion of DNA-binding peptide to Tbr DNA pol results in lower specificity of the DyNAmo kit which is evident in false positive results for no template control (NTC) reactions. Chemically modified polymerase produces delayed Cts and lower signal strength compared to AccuStart™ Taq. Cycling conditions: Finnzymes: 95°;C, 7 min followed by 40 cycles of 95°C, 10s; 60°C, 20s PerfeCTa™ SYBR Green FastMix: 95°;C, 20s followed by 40 cycles of 95°C, 1s; 60°C, 20s


Comparison to Takara, ADAR

PageW - Copy of Graph ADAR - ADAR FastMix vs Takara*

PerfeCTa SYBR Green FastMix Comparison to SYBR PreMix Ex Taq
RNA-specific adenosine deaminase (ADAR) was amplified from log-fold dilutions of total HeLa cell cDNA (100 ng to 10 pg) using PerfeCTa™ SYBR Green FastMix or SYBR PreMix Ex Taq™ (Takara) according to each manufacturers protocol. Averaged plots for quadruplicate reactions for each input quantity are shown. PerfeCTa™ SYBR Green FastMix produces higher fluorescent signal and detection of equal target amounts at earlier Cts. Cycling conditions for both kits: 95°C, 20s followed by 40 cycles of 95°C, 1s; 60°C, 20s.

Comparison to SYBR GreenER, ADAR

PageW - Graph Page 1 - Data 1

PerfeCTa SYBR Green FastMix comparison to SYBR GreenER qPCR SuperMix
RNA-specific adenosine deaminase (ADAR) was amplified from log-fold dilutions of total HeLa cell cDNA (100 ng to 1 pg) using PerfeCTa™ SYBR Green FastMix or the SYBR GreenER qPCR SuperMix (Invitrogen) according to each manufacturers protocol. Averaged plots for quadruplicate reactions for each input quantity are shown. Replicate CT values are shown on the standard curve (Panel A, inset). Cycling conditions: Invitrogenn: 95°C, 10 min followed by 40 cycles of 95°C, 10s; 60°C, 60s; PerfeCTa™ SYBR Green FastMix: 95°C, 20s followed by 40 cycles of 95°C, 1s; 60°C, 20s. PerfeCTa SYBR Green FastMix amplified the ADAR gene with higher efficiency and greater sensitivity. All replicate reactions for SYBR GreenER qPCR SuperMix failed to amplify ADAR from 1 pg of cDNA.

Contents

  • 2X reaction buffer containing optimized concentrations of MgCl2, dNTPs (dATP, dCTP, dGTP, dTTP), AccuFast Taq DNA Polymerase, SYBR Green I dye, 20 nM fluorescein, and stabilizers.

Storage conditions

PerfeCTa SYBR Green FastMix is stable for 1 year when stored in a constant temperature freezer at -20°C, protected from light. For convenience, it may be stored unfrozen at 4°C for up to 6 months. After thawing, mix thoroughly before using. Stabilized reagent demonstrates no functional loss of performance after 20 freeze-thaw cycles or 2 months at 20°C.

Resources

Ordering

Cat# 95071-250 250 x 20-µl rxns (2 x 1.25 ml)
Cat# 95071-012 1250 x 20-µl rxns (10 x 1.25 ml)
Cat# 95071-05K 5000 x 20-µl rxns (1 x 50 ml)