QIAGEN OneStep RT-PCR Kit (100)

A one-step reverse transcription PCR system from QIAGEN carries out reverse transcription and PCR in a single tube from an RNA template. The enzyme mix combines HotStarTaq DNA Polymerase with Omniscript and Sensiscript Reverse Transcriptases, transcribing RNA amounts from 1 pg up to 2 µg.

The OneStep RT-PCR Buffer contains additives that prevent inhibition of PCR by the reverse transcriptases and provides 2.5 mM MgCl2 at 1x working concentration. The buffer's K+ and NH4+ ions support specific primer annealing across a wide temperature range. Q-Solution, also included, modifies nucleic acid melting behaviour to assist amplification of GC-rich templates. After reverse transcription at 50°C, an incubation of 15 minutes at 95°C activates HotStarTaq DNA Polymerase and inactivates the reverse transcriptases, reducing non-specific products such as primer dimers.

Gene-specific primers are used at a recommended final concentration of 0.6 µM; random oligomers or oligo-dT primers are not recommended. Typical thermal cycling uses 25–40 cycles of denaturation at 95°C, annealing at 50–68°C, and extension at 72°C, with a 40-second extension sufficing for products up to 300 bp. Reactions are endpoint, single-plex, and hot-start activated. The kit can be applied to virus detection, gene-expression analysis, and molecular diagnostics research using total RNA, mRNA, or viral RNA. It is intended for molecular biology research use only and is not intended for the diagnosis, prevention, or treatment of disease.