OneStep RT-qPCR

MasterMixes

Reverse transcription & qRT-PCR in one tube
Efficient cDNA synthesis ensured by the thermostable Reverse Transcriptase & advanced RNase Inhibitor mix
Works with high GC & high AT-rich templates
Rapid extension, early Ct detections
Very high sensitivity

Relative gene expression analysis
Absolute quantification of any RNA template (mRNA, total RNA, viral RNA), low copy number genes
qRT-PCR assays based on specific probes: including TaqMan®, Molecular Beacons, Scorpions™ Probes

2X master mix for 500 reaction per 20µl reaction volume
Optimized with Hot Start Polymerase, Thermostable, and extremely active Reverse Transcriptase & advanced RNase Inhibitor allowing for a single step one tube qRT-PCR.
Highest sensitivity and specificity under both standard and fast qPCR cycling conditions.
Excellent performance on both AT/GC rich templates and guaranty rapid extension with early Ct values with minimum or no optimization.
Supplied with PCR Water to guaranty the best performance.
To cover all RT-PCR instruments, the OneStep qRT-PCR Probe-based MasterMixes are available in three versions – without ROX, with low or high ROX concentration. See the ROX concentration table below to know which ROX concentration suits your instrument

Tips for successful workflow

Use special primer selection programs for good planning.
Work with amplicons in a range of 80-200, max 400 bp.
Take typical measures to prevent PCR cross over contamination, keep your bench clean, wear gloves, use sterile tubes and filter pipet tips.
Run reactions in triplets; include a no-template control, no RT Mix control and positive control in parallel.
Thaw and keep reagents on ice. Mix well before use.
Do not perform annealing/extension for more than 30 seconds and do not use lower than 60 °C temperature for this step.

To Prepare a 20 μl reaction, mix the following:

Reverse Primer: 100 - 400 nM final concentration.
Forward Primer: 100 - 400 nM final concentration.
Total RNA template: 1pg to 1 μg (or mRNA template; >0.01 pg)
Add PCR Water to 10 μl volume
OneStep qRT-PCR MMix (2X): 10 μl
RT3 Mix (20X) 1-2 μl (2 μl improve Ct, but primer dimers may appear)
Mix gently, avoid bubbles. Place into the instrument
- If Probe based MasterMix is used set the instrument channels according to the probes specifications
- If Green MasterMix is used, set the instrument onto SYBR® Green or FAM channel

PCR Program

ROX