Titrate the siRNA: Use different concentrations of siRNA and consult the manufacturer’s instructions, generally a concentration of 5–100 nM is used. Use the lowest working concentration. Monitor RNA and protein level: Successful mRNA silencing without a corresponding decrease in protein level suggests...
0.25 nmol of each siRNA is provided dried-down in the wells of barcoded 96- or 384-well plates. This is sufficient for 100-500+ transfections, depending on siRNA working concentration and reaction volume. Column 12 of each 96-well plate and columns 23 and 24 in...
0.25 nmol of each siRNA is provided dried-down in the wells of barcoded 96- or 384-well plates. This is sufficient for 100-500+ transfections, depending on siRNA working concentration and reaction volume. Column 12 of each 96-well plate and columns 23 and 24 in ...
6. Validation of siRNA data Titrate the siRNA:Use different concentrations of siRNA and consult the manufacturer’s instructions, generally a concentration of 5–100 nM is used. Use the lowest working concentration. Monitor RNA and protein level:Successful mRNA silencing without a corresponding decreas...
There are many influencing factors for transfection efficiency, among them the cell line, siRNA concentration and its ratio to the transfection reagent, cell confluence during transfec- tion, incubation time and media composition all need to be taken intoconsideration. In this Application Note we ...
calculated relative to untreated cells.WhilePepMute™ and Dharmafect™ 4 reagents delivered significant gene silencing from 1.0 nM of renilla luciferase siRNA,Lipofectamine™ RNAiMAX gave good knockdown only after 20 nM while enhanced gene expression at low concentration of siRNA (0.5 and 1.0 nM ...
Optimal siRNA concentration 30 nMDiluent for siRNA and GenMute™ Reagent 1xGenMute™ Transfection BufferIncubation Time to Form GenMute™/siRNA Complex ~15 minutes at RTMaximal Silencing Efficiency 48 hoursTransfection Results:Reporter Gene Renilla Luciferase Silencing Efficiency (% ) 90%Storage:...
Summary of Optimal Transfection Conditions: Cell culture medium Confluency of cell on the day of transfection Optimal siRNA concentration Diluent for siRNA and GenMute™Reagent Incubation Time to Form GenMute™/siRNA Complex Maximal Silencing Efficiency ...
(50 ul total volume of assembled labeling mix or 5 ug of 21 mer duplex siRNA) for the labeling procedure. After performing the labeling reaction, ethanol precipitation and re-suspension, I pool the aliquots before determining labeled RNA concentrations using Ambion’s online concentration calculator...
The supernatant was collected continuously for 5 days, and the siRNA concentration was calculated using a Nanodrop spectrometer (Thermo Fisher Scientific). BV2 cell culture, transfection and cell viability assay The BV2 immortalized murine microglial cell line was received from Dr. Eun-Hye Joe of ...