Universal Fast One-Step RT-qPCR Probe Kit (U⁺) II is designed for one-step real-time RT-PCR with probe-based detection. The kit enables completion of 42 amplification cycles within 40 minutes. The included 25× One-Step Enzyme Mix (U⁺) II contains heat-labile Uracil N-Glycosylase (UNG), which effectively degrades carryover contamination from previous amplifications to minimize false positives.

5× One-Step buffer (U⁺) II: 4 μL
25× One-Step Enzyme mix (U⁺) II: 0.8 μL
Primer Forward: 0.1-1.0 μM
Primer Reverse: 0.1-1.0 μM
TaqMan Probe: 0.05-0.2 μM
RNA Target: X μL
RNase Free Water: Up to 20 μL

Standard Procedure
Reverse Transcription: 55°C for 10:00 (1 cycle)
Initial Denaturation: 95°C for 0:30 (1 cycle)
Denaturation: 95°C for 0:10 (45 cycles)
Annealing & Extension: 60°C for 0:30 (45 cycles)
Fast Procedure
Reverse Transcription: 55°C for 3:00 (1 cycle)
Initial Denaturation: 95°C for 0:30 (1 cycle)
Denaturation: 95°C for 0:05 (42 cycles)
Annealing & Extension: 60°C for 0:10 (42 cycles)
Reverse transcription temperature ranges from 50°C to 60°C. Increasing temperature improves amplification efficiency for complex or high-GC templates. Adjust annealing temperature based on primer Tm. Use shortest possible signal acquisition time per PCR instrument specifications.

Ship with ice packs. Store at -20°C. Avoid repeated freeze-thaw cycles. Product is stable for 12 months under recommended storage conditions.