After a RNA extraction, 1% agarose gel is normally performed to verify the quality/integrity of the RNA. It seems like RNA extraction is fine as far as two bands appear (with no smear), corresponding to 18S and 28S RNA, regardless their molecular weight. But I have found in different RNA extractions (same samples and same protocol), different molecular weights for the two bands, putatively corresponding to 18S and 28S. Any suggestion of what's going on? Knowing the actual molecular weight of 18S an 28S RNA could help, but I couldn't find it anywhere.
three types of ribosomal RNA are present in a plant cell, mitochondrial, chloroplastic and nuclear. This is a first possible source of variation. Nuclear rRNA may be differently methylated (and perhaps other modifications) in different tissues or at differing stages of development. This is a second source of variation. Your step of denaturation before deposing the samples may not be exactly the same for all samples, a third possible source of variation!
What you are seeing might be the bands of 23S and 16S rRNA of the plastids. If you are working with green leaves, you might be extracting more of these two from the chloroplasts than of the nuclear 28S and 18S.
Even if you do RIN anaylsis on a bioanalyzer you get lower values on green plant samples, due to chloroplastic rRNA. After talking to an Agilent agent I found there is nor sure assay to find accurate RIN numbers for green plants.
three types of ribosomal RNA are present in a plant cell, mitochondrial, chloroplastic and nuclear. This is a first possible source of variation. Nuclear rRNA may be differently methylated (and perhaps other modifications) in different tissues or at differing stages of development. This is a second source of variation. Your step of denaturation before deposing the samples may not be exactly the same for all samples, a third possible source of variation!
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