I just started working on miRNA and I still have a lot of doubts about them. I'm going to validate some miRNA expression using TaqMan miRNA Assay (Life) in serum from mice and human. My question is about the controls. For mice, I bought some snRNA and snoRNA. But for human, I don't know what to use. I got one spike-in, but if I understand it right, it is used just to monitor sample preparation. So, to normalize my data, do I have to use another control, since snoRNA and snRNA are not usually present in serum? Like U6 or SNORD RNAs?
Dear Fargoso,
for extracellular miRNAs U6 or SNORED will not work properly as they are not abundantly available in extracellular condition (they are particularly good for cellular control). if you use panel of miRNAs (more than 450) global normalization will give you the best result. if you use selected number of miRNAs (8-10) then miR-103, miR-191 or miR-16 might be helpful.
hope this will work. if you have further query please do not hesitate to contact me.
Dear Mariana Franco Fragoso
I suggestive you following tips.
1. There is no endogenous control to normalize serum miRNA expression.
2. While isolating RNA from Serum samples add synthetic cel-miR-39 ( Qaigen). then proceed isolation.
3. After qPCR , use cel-miR-39 as endogenous control for normalization.
Please find attached supporting article
For further clarification let me know.
with regards
pp
- Badges
- Science topic
- Similar topics
- Biological Science
- Biochemistry
- Biomolecules
- Nucleic Acids
- RNA
- Small RNA
- microRNA