I started using "RNA storage solution (1mM sodium citrate) from THermoFisher to dissolve RNA pellets and store in -20 or -80.
However, there is no amplification in the Q-PCR. it is frustrating and I am not sure if the solution affects the RNA, and if no do I have to change the RNA input?
I use 2 mcg RNA to make cDNA with high capacity superscriptII RT-kit (20 ul). but no luck so far in q-PCR?
Hi Kamelia,
sodium citrate does not have any impact on RNA downstream applications as far as I know. However, did you try water as a solvent and then performed RT? As for your PCR, did you have successfully used your assay protocol on different input samples (i.e. RNA from HeLa)? Any housekeeping genes as positive control? If you use 2 µg RNA try using less and perform a regular PCR with subsequent gel seperation.
Best regards
Soner
Citrate is a chelator and it may mess up the Mg2+ concentration required by reverse transcriptase and DNA polymerase. However, if the final concentration of citrate is, say an order of magnitude lower than the Mg2+ concentration, it shouldn't affect the reaction too much
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