I have isolated RNA from keratinocyte cells using Trireagent from SigmaAldrich. With varying concentration, low 260/230, and a spectra that matches phenol contamination, I suspect.... phenol.
The RNA pellet was re-suspended in H20. Is there a simple lossless way to remove the phenol contamination? Also, could I still perform a reverse transcriptase-qPCR with the contamination or does the phenol hinder downstream application?
firstly when isolating RNA take care while taking out supernatent containing RNA, do not take below the precipitate layer, nor disturb.
second, Increase the Ethanol wash frequency, generally one wash is enough but you can wash twice.
Do an extraction with an equal volume of 2-butanol (add 1 volume of 2-buOH, vortex, spin for 1 min, discard the upper organic phase and then reprecipitate with NaAc/EtOH). Phenol will partition into the organic phase, RNA will remain in the aqueous phase.
Beats using the expensive -though very effective- columns of Qiagen et al.
Hope it helps!
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