Guidelines for transfection
miRNA mimic or inhibitor transfection
Plasmid DNA and miRNA mimic/inhibitor cotransfection
Optimizing miRNA transfection
The amount of miRNA mimic/inhibitor needed to efficiently downregulate a target gene or inhibit miRNA function can vary greatly, depending on the miRNA, the cell line, and the chosen analysis method. To determine the concentration that provides optimal results, optimization experiments using varying mimic/inhibitor concentrations should be performed.
miRNA mimics can inhibit target protein expression at a final concentration as low as 0.5 nM. However, a higher concentration may be required, especially if performing downstream analysis at the protein level. miRNA inhibitors have been shown to inhibit miRNA function at a concentration of 50 nM. Lower inhibitor concentrations may also be effective.
In addition to optimization of concentration, time-course experiments may also be necessary to determine the optimal time after transfection for analysis of results. miRNA mimic or inhibitor effects often do not lead to an immediate change in transcript or protein levels.