How should I choose between a one-step RT-qPCR or two-step RT-qPCR approach?

Two methods are available for quantifying RNA samples: one-step and two-step RT-qPCR. In both cases, RNA is first reverse transcribed into cDNA, which is then used as the template for qPCR amplification. Reverse transcription can be performed separately from qPCR (i.e., two-step RT-qPCR) or directly in the qPCR mix (i.e., one-step RT-qPCR). One-step workflows are commonly favored in molecular diagnostic assays, where sample inputs may be limiting or numerous samples are examined. Two-step RT-qPCR is preferred when multiple interrogations will be made of the same starting material or where archiving of cDNA may be required. For more details, please refer to “Choice of One-Step RT-qPCR or Two-Step RT-qPCR”.

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