Dear all,
I am still facing a lot of troubles with my RNA extraction by TRIzol. Despite my attention in avoiding any kind of contaminations (especially with the interphase/organic phase), removing all the ethanol residues etc. 260/230-260/280 ratios are really bad. This damages the quality of my qPCR experiments. It's been 5-6 months since I've started to have this kinds of problems. Previously, I extracted my RNA from frozen pieces of tissues, now I only work on cryostate cuts conditioned by Tragacanth gum.
What are your ratios? Different people have different levels of contamination they are willing to tolerate. I've used Trizol purification with Phase-Lock tubes, and I've often gotten very good results in RT-qPCR despite a high 230nm absorbance (often indicative of phenolate or quanidine contamination).
In some instances (e.g. in TE buffer) ethanol contamination itself doesn't have a meaningful impact on absorbance spectra.
You might be able to reduce the level of such contamination with an additional precipitation step, or via Chisom washing, or via purification using an RNeasy column.
Agree with @ Harald Kühnel
you should check the quality of the water and tubes as well as take into consideration the absence of pollution
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