I have studied miRNA expression by calculating 2-ddCt . After I have performed qRT-PCR of target and reference miRNAs (hsa-miR-16-5p), I calculate delta Ct by using this formular " Ct(target miRNA) - Ct (hsa-miR-16-56). I find that some values present negative results such as "Ct(target) - Ct (hsa-miR-16-5p: reference) = -2.12". I would like to discuss with you in some points as follows:
1. Should I change reference gene?
2. Can these results occur and Can I use these results for further data analysis?
Thank you very much.
Dr. Methee
Dear Sriprapun,
In your experiential situation, the Ct value of the target gene is much earlier than the reference gene and it is very much possible. Having a lesser Ct value for GOI compared to Endogenous Control means that your gene of interest is expressed at a higher level than Endogenous Control. Definitely, You can go for the data analysis.
With regards,
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