What is an extra band in gel electrophoresis of total RNA?

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Andrei S. Babenka

hi, for me 28S between 1,5-2.0 kb, 18S between 700 and 800 bp. On the top it could be another fractions....may be some DNA, may be rRNA complexes. You also can find some bands with a low intensity in 250-300 bp and oth. It could be 5.8S rRNA. For me the results is quiete good but it would be better to use Bioanalyzer or something else with capillary electrophoresis.

Alireza Mordadi

I wasn't sure but now with Andrei,s answer I am sure. He's right.

You can check it for DNA with cheaper method, use primer and conventional PCR.

Regards

Motahareh Nadimi

Thanks Andrei and Alireza for all your suggestions.

you mean the top band is DNA not 28sRNA? 28s rRNA isn't ~(5 kb)?

Yordan Muhovski

Hi,

I disagree with the comments. For me the upper band correspond perfectly to human 28s rRNA which is around 5kb and the second band is 18s rRNA or around 1.9. The last band of your DNA ladder, if correct, is 1.5kb (according to your attached picture). The last RNA band around 700 bp could be some specific PBMC RNAs. Well, the other point to mention is that you used DNA ladder to estimate RNA size which is moreless correct but ss nucleic acids migrate slightly differnt from ds DNA in your ladder. Anyway I don't see DNA contamination in your samples. But using Bioanalyzer 2100 is more accurate.

No.

The top bond is 28s rena ~5.0 kb.

Our point is smear form and other bands, as mentioned by Andrei is most probably RNA and mentioned by Yordan in other way.

And absolutely yes, the methods mentioned by both is perfect by as I am familiar with situation it's better use conventional PCR.

Adriano Costa de Alcantara

Check the data from promega at the following website: https://www.promega.com.br/resources/pubhub/methods-of-rna-quality-assessment/

At this other website you will be ale to see a gel done by the company showing the result of RNA extraction showing bands as yours (Unfortunately, there is no marker presented for us to check if it is the same size of yours. Anyway, I think it is).

https://www.sigmaaldrich.com/catalog/product/sigma/rtn70?lang=pt®ion=BR

The only thing I would say as an extra advice for you is when you finish your extraction and before you freeze your sample, take an aliquot to do your quantification by whatever method you decide to use. Do not freeze/thaw your sample because RNA is very unstable and degradable. Ok?

All the best.