PerfeCTa SYBR Green FastMix, ROX

PerfeCTa® SYBR® Green FastMix, ROX™ is a 2X concentrated, ready-to-use reaction cocktail that contains all components, except primers and template for real-time quantitative PCR on Applied Biosystems 7000, 7300, 7700, or 7900 instruments. 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.

Performance Data

Comparison to Takara, ADAR

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

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.

Real-Time PCR Instrument Compatibility

• Applied Biosystems 5700
• Applied Biosystems 7000
• Applied Biosystems 7300
• Applied Biosystems 7700
• Applied Biosystems 7900
• Applied Biosystems 7900HT
• Applied Biosystems 7900 HT Fast
• Applied Biosystems StepOne™
• Applied Biosystems StepOnePlus™

Contents

• Single-tube, 2X concentrated reagent containing:
  • Reaction buffer with optimized concentrations of molecular-grade MgCl2, dATP, dCTP, dGTP, and dTTP.
  • AccuStart II Taq DNA Polymerase
  • SYBR Green I dye
  • Inert AccuVue dye
  • Proprietary enzyme stabilizers and performance-enhancing additives.
  • Titrated reference dye (if applicable)

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

• 95073 PerfeCTa SYBR Green FastMix, ROX Protocol
• 95073 PerfeCTa SYBR Green FastMix, ROX MSDS
• SYBR SuperMixes and FastMixes Product Brochure

Ordering

Cat# 95073-250       250 x 20-µl rxns (2 x 1.25 ml)

Cat# 95073-012       1250 x 20-µl rxns (10 x 1.25 ml)

Cat# 95073-05K      5000 x 20-µl rxns (1 x 50 ml)