Actually I am working with primary B cells and I need to isolate the RNA with high concentration and purity , I tried a lot of kits but still the RNA concentration is low and I could not see the isolated RNA in gel electrophresis ?
Hello Sawsan,
RNA detection in agarose gels varies depending on the denaturant, stain, and image acquisition conditions used.
Using Ethidium bromide, you can detect 30 ng of native RNA and 150 ng of Glyoxal RNA.
Not sure which kits you are using but good quality RNA can be isolated in good amounts (~5 ug from 1E6 cells) using Trizol reagent. If you have Trizol, please go through the following protocol.
https://tools.thermofisher.com/content/sfs/manuals/trizol_reagent.pdf
Hello
Thanks Ali for quick response , I used the Ribo pure kit ; Trizol reagent and I measured the RNA concentration with nano drop also I got 80 ng / ul from 20 million and 40ng from 2 million , besides to I prepared 1.5 agarose gel in TAE ,every thing as molecular grade and I used ethidium bromide
but I could not see RNA in gel
Well I am not a fan of nano drop; I use Invitrogen's Qubit fluorometer instead. If you didn't see nothing in the gel then perhaps you dont have it there, not in good concentration at least. Is this for the first time you are isolating RNA or you have prior experience? Your RNA yield appear to be very low. Are you sure the RNA is not getting degraded?
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