Variability of the reverse transcription step: Practical implications
Bustin S., Dhillon HS., Kirvell S., Greenwood C., Parker M., Shipley GL., Nolan T.
Methods: We investigated the reproducibility of the RT step with commercial reverse transcriptases and RNA samples of variable quality and concentration. We quantified several mRNA targets with either singleplex SYBR Green I or dualplex probe-based reverse transcription real-time quantitative PCR (RT-qPCR), with the latter used to calculate the correlation between quantification cycles (Cqs) of mRNA targets amplified in the same realtime quantitative PCR (qPCR) assay.