I am trying to extract RNA from frozen liver tissue samples for gene expression analysis. The ratio look good on Nanodrop but when i proceed it on gel it shows degradation (smears).
what does it mean ?
Can someone help ? It will be a great favor.
Nanodrop will just tell you the concentration, but not the length or quality of nucleotides. Degraded RNA will look "the same" as intact RNA on a spectrophotometer.
It's normal to see a bit of a smear from the mRNAs, but the rRNAs should show up as distinct bands.
Ask a colleague to take a took for you.
As Katie said, degraded "smear" RNA looks the same as intact RNA in Nanodrop. If you want to prevent RNA degradation from frozen livers, I recommend using RNAlater. If you are using an extraction method such as TRIzol, it is better to use a column method such as RNeasy. And in some cases, using 50% ethanol (instead of 70% ethanol) may increase RNA yields from liver samples.
Yohei Kono Katie A S Burnette thankyou so much . I will try your suggested methods.
All the suggestions are good, I will also suggest you don't mind the smear bands and go on with your Reverse transcription reaction for gene expression study (if it's 2 Step) , since you already have a good nanodrop readings.
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