I carried out total RNA non-denaturing agarose gel electrophoresis from 2 human cancer cells. I am not sure what to make of the results as I am finding conflicting statements. Are the first and second bands 28S and 18S? Or is the first band genomic DNA contamination? Also the ladder is a DNA ladder. Please find the gel image attached to the question.
Hi, Fatima Farouk
Initially, next time, use an RNA ladder to evaluate RNA samples. Ok? (Of course, If you did not have it there, it was a nice idea to at least put some ladder).
Anyway, Looking at the picture, I could notice the first band (upper at the gel picture) is still inside the well. So, usually, I would think it as a gDNA. Ok?
Looking at the next bands, it seems to me a nice result (too much sample of U87 MG) and nice amount of MCF7. One thing, it is probably good enough for RT-qPCR and other downstream applications but the ratio 28S:18S are not twice. So, maybe, your cells were already dying (old or treatment is killing them or your time to extract/freezing are doing it) and your RNA is getting degraded. Ok?
Hope it helps.
Hello all, thank you for your inputs. I did a nanodrop of the RNA before gel electrophoresis, and I had very good purity ratios, so I went ahead to make cDNA, did both benchtop PCR and qPCR. However, I need to include the RNA gel electrophoresis results, and I am not sure what to make of the gel image. What are the expected base-pair lengths of the 28S, 18S and 5S rRNAs? I would really appreciate a reliable article, because I am finding conflicting statements.
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